3-(heteroaryl) alanine derivatives-inhibitors of leukocyte adhesion mediated by VLA-4

ABSTRACT

Disclosed are certain 3-(heteroaryl)alanine derivatives which bind VLA-4 and inhibit leukocyte adhesion mediated by VLA-4. Such compounds are useful in the treatment of inflammatory diseases in a mammalian patient, e.g., human, such as asthma, Alzheimer&#39;s disease, atherosclerosis, AIDS dementia, diabetes, inflammatory bowel disease, rheumatoid arthritis, tissue transplantation, tumor metastasis and myocardial ischemia. The compounds can also be administered for the treatment of inflammatory brain diseases such as multiple sclerosis.

CROSS REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of U.S. ProvisionalApplication Ser. No. 60/220,131, filed Jul. 21, 2000, which isincorporated by reference in its entirety.

BACKGROUND OF THE INVENTION

[0002] 1. Field of the Invention

[0003] This invention relates to certain 3-(heteroaryl)alaninederivatives which inhibit leukocyte adhesion and, in particular,leukocyte adhesion mediated by VLA-4.

[0004] References

[0005] The following publications, patents and patent applications arecited in this application as superscript numbers:

[0006] 1 Hemler and Takada, European Patent Application Publication No.330,506, published Aug. 30, 1989

[0007] 2 Elices, et al., Cell, 60:577-584 (1990)

[0008] 3 Springer, Nature, 346:425-434 (1990)

[0009] 4 Osborn, Cell, 62:3-6 (1990)

[0010] 5 Vedder, et al., Surgery, 106:509 (1989)

[0011] 6 Pretolani, et al., J. Exp. Med., 180:795 (1994)

[0012] 7 Abraham, et al., J. Clin. Invest., 9:776 (1994)

[0013] 8 Mulligan, et al., J. Immunology, 150:2407 (1993)

[0014] 9 Cybulsky, et al., Science, 251:788 (1991)

[0015] 10 Li, et al., Arterioscler. Thromb., 13:197 (1993)

[0016] 11 Sasseville, et al., Am. J. Path., 144:27 (1994)

[0017] 12 Yang, et al., Proc. Nat. Acad. Science (USA), 90:10494 (1993)

[0018] 13 Burkly, et al., Diabetes, 43:529 (1994)

[0019] 14 Baron, et al., J. Clin. Invest., 93:1700 (1994)

[0020] 15 Hamann, et al., J. Immunology, 152:3238 (1994)

[0021] 16 Yednock, et al., Nature, 356:63 (1992)

[0022] 17 Baron, et al., J. Exp. Med., 177:57 (1993)

[0023] 18 van Dinther-Janssen, et al., J. Immunology, 147:4207 (1991)

[0024] 19 van Dinther-Janssen, et al., Annals. Rheumatic Dis., 52:672(1993)

[0025] 20 Elices, et al., J. Clin. Invest., 93:405 (1994)

[0026] 21 Postigo, et al., J. Clin. Invest., 89:1445 (1991)

[0027] 22 Paul, et al., Transpl. Proceed., 25:813 (1993)

[0028] 23 Okarhara, et al., Can. Res., 54:3233 (1994)

[0029] 24 Paavonen, et al., Int. J. Can., 58:298 (1994)

[0030] 25 Schadendorf, et al., J. Path., 170:429 (1993)

[0031] 26 Bao, et al., Diff., 52:239 (1993)

[0032] 27 Lauri, et al., British J. Cancer, 68:862 (1993)

[0033] 28 Kawaguchi, et al., Japanese J. Cancer Res., 83:1304 (1992)

[0034] 29 Kogan, et al., U.S. Pat. No. 5,510,332, issued Apr. 23, 1996

[0035] 30 International Patent Appl. Publication No. WO 96/01644

[0036] All of the above publications, patents and patent applicationsare herein incorporated by reference in their entirety to the sameextent as if each individual publication, patent or patent applicationwas specifically and individually indicated to be incorporated byreference in its entirety.

[0037] 2. State of the Art

[0038] VLA-4 (also referred to as α₄β₁ integrin and CD49d/CD29), firstidentified by Hemler and Takada¹ is a member of the β1 integrin familyof cell surface receptors, each of which comprises two subunits, an αchain and a β chain. VLA-4 contains an α4 chain and a β1 chain. Thereare at least nine β1 integrins, all sharing the same β1 chain and eachhaving a distinct α chain. These nine receptors all bind a differentcomplement of the various cell matrix molecules, such as fibronectin,laminin, and collagen. VLA-4, for example, binds to fibronectin. VLA-4also binds non-matrix molecules that are expressed by endothelial andother cells. These non-matrix molecules include VCAM-1, which isexpressed on cytokine-activated human umbilical vein endothelial cellsin culture. Distinct epitopes of VLA-4 are responsible for thefibronectin and VCAM-1 binding activities and each activity has beenshown to be inhibited independently.²

[0039] Intercellular adhesion mediated by VLA-4 and other cell surfacereceptors is associated with a number of inflammatory responses. At thesite of an injury or other inflammatory stimulus, activated vascularendothelial cells express molecules that are adhesive for leukocytes.The mechanics of leukocyte adhesion to endothelial cells involves, inpart, the recognition and binding of cell surface receptors onleukocytes to the corresponding cell surface molecules on endothelialcells. Once bound, the leukocytes migrate across the blood vessel wallto enter the injured site and release chemical mediators to combatinfection. For reviews of adhesion receptors of the immune system, see,for example, Springer³ and Osborn⁴.

[0040] Inflammatory brain disorders, such as experimental autoimmuneencephalomyelitis (EAE), multiple sclerosis (MS) and meningitis, areexamples of central nervous system disorders in which theendothelium/leukocyte adhesion mechanism results in destruction tootherwise healthy brain tissue. Large numbers of leukocytes migrateacross the blood brain barrier (BBB) in subjects with these inflammatorydiseases. The leukocytes release toxic mediators that cause extensivetissue damage resulting in impaired nerve conduction and paralysis.

[0041] In other organ systems, tissue damage also occurs via an adhesionmechanism resulting in migration or activation of leukocytes. Forexample, it has been shown that the initial insult following myocardialischemia to heart tissue can be further complicated by leukocyte entryto the injured tissue causing still further insult (Vedder et al.⁵).Other inflammatory or medical conditions mediated by an adhesionmechanism include, by way of example, asthma⁶⁻⁸, Alzheimer's disease,atherosclerosis⁹⁻¹⁰, AIDS dementia¹¹, diabetes¹²⁻¹⁴ (including acutejuvenile onset diabetes), inflammatory bowel disease¹⁵ (includingulcerative colitis and Crohn's disease), multiple sclerosis¹⁶⁻¹⁷,rheumatoid arthritis¹⁸⁻²¹, tissue transplantation²², tumormetastasis²³⁻²⁸, meningitis, encephalitis, stroke, and other cerebraltraumas, nephritis, retinitis, atopic dermatitis, psoriasis, myocardialischemia and acute leukocyte-mediated lung injury such as that whichoccurs in adult respiratory distress syndrome.

[0042] In view of the above, assays for determining the VLA-4 level in abiological sample containing VLA-4 would be useful, for example, todiagnosis VLA-4 mediated conditions. Additionally, despite theseadvances in the understanding of leukocyte adhesion, the art has onlyrecently addressed the use of inhibitors of adhesion in the treatment ofinflammatory brain diseases and other inflammatory conditions^(29,30).The present invention addresses these and other needs.

SUMMARY OF THE INVENTION

[0043] This invention provides certain 3-(heteroaryl)alanine derivativeswhich bind to VLA-4. Such compounds can be used, for example, to assayfor the presence of VLA-4 in a sample and in pharmaceutical compositionsto inhibit cellular adhesion mediated by VLA-4, for example, binding ofVCAM-1 to VLA-4. The compounds of this invention have a binding affinityto VLA-4 as expressed by an IC₅₀ of about 15 μM or less (as measuredusing the procedures described in Example A below).

[0044] In particular, this invention is based on the discovery that thepresence of a nitrogen containing heteroaryl group at the 3-position ofthese alanine derivatives provides significant advantages vis-a-vis arylor other heteroaryl groups at this position in the molecule.

[0045] Accordingly, in one of its composition aspects, this invention isdirected to a compound of Formula (I):

[0046] wherein:

[0047] A is selected from the group consisting of aryl, heteroaryl,cycloalkyl, and heterocyclic group wherein said aryl, heteroaryl,cycloalkyl, or heterocyclic group is optionally substituted, on any ringatom capable of substitution, with 1-3 substituents selected from thegroup consisting of alkyl, substituted alkyl, alkoxy, substitutedalkoxy, acyl, acylamino, thiocarbonylamino, acyloxy, amino, substitutedamino, amidino, alkyl amidino, thioamidino, aminoacyl,aminocarbonylamino, aminothiocarbonylamino, aminocarbonyloxy, aryl,substituted aryl, aryloxy, substituted aryloxy, aryloxyaryl, substitutedaryloxyaryl, cyano, halogen, hydroxyl, nitro, oxo, carboxyl, cycloalkyl,substituted cycloalkyl, guanidino, guanidinosulfone, thiol, thioalkyl,substituted thioalkyl, thioaryl, substituted thioaryl, thiocycloalkyl,substituted thiocycloalkyl, thioheteroaryl, substituted thioheteroaryl,thioheterocyclic, substituted thioheterocyclic, heteroaryl, substitutedheteroaryl, heterocyclic, substituted heterocyclic, cycloalkoxy,substituted cycloalkoxy, heteroaryloxy, substituted heteroaryloxy,heterocyclyloxy, substituted heterocyclyloxy, oxycarbonylamino,oxythiocarbonylamino, —OS(O)₂-alkyl, —OS(O)₂-substituted alkyl,—OS(O)₂-aryl, —OS(O)₂-substituted aryl, —OS(O)₂-heteroaryl,—OS(O)₂-substituted heteroaryl, —OS(O)₂-heterocyclic,—OS(O)₂-substituted heterocyclic, —OSO₂—NRR where each R isindependently hydrogen or alkyl, —NRS(O)₂-alkyl, —NRS(O)₂-substitutedalkyl, —NRS(O)₂-aryl, —NRS(O)₂-substituted aryl, —NRS(O)₂-heteroaryl,—NRS(O)₂-substituted heteroaryl, —NRS(O)₂-heterocyclic,—NRS(O)₂-substituted heterocyclic, —NRS(O)₂—NR-alkyl,—NRS(O)₂—NR-substituted alkyl, —NRS(O)₂—NR-aryl, —NRS(O)₂—NR-substitutedaryl, —NRS(O)₂—NR-heteroaryl, —NRS(O)₂—NR-substituted heteroaryl, —NRS(O)₂—NR-heterocyclic, —NRS(O)₂—NR-substituted heterocyclic where R ishydrogen or alkyl, —N[S(O)₂—R′]₂ and —N[S(O)₂—NR′]₂ where each R′ isindependently selected from the group consisting of alkyl, substitutedalkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl,heterocyclic and substituted heterocyclic;

[0048] HetAr is a nitrogen containing heteroaryl or a nitrogencontaining substituted heteroaryl group;

[0049] Alk is an alkylene group of 1 to 4 carbons;

[0050] m is 0 or 1;

[0051] R¹ is selected from the group consisting of hydrogen, alkyl,substituted alkyl, alkenyl, substituted alkenyl, cycloalkyl, substitutedcycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substitutedaryl, heteroaryl, substituted heteroaryl, heterocyclic and substitutedheterocyclic;

[0052] X is selected from the group consisting of hydroxyl, alkoxy,substituted alkoxy, alkenoxy, substituted alkenoxy, cycloalkoxy,substituted cycloalkoxy, cycloalkenoxy, substituted cycloalkenoxy,aryloxy, substituted aryloxy, heteroaryloxy, substituted heteroaryloxy,heterocyclyloxy, substituted heterocyclyloxy and —NR″R″ where each R″ isindependently selected from the group consisting of hydrogen, alkyl,substituted alkyl, alkenyl, substituted alkenyl, cycloalkyl, substitutedcycloalkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl,heterocyclic and substituted heterocyclic;

[0053] and enantiomers, diasteromers and pharmaceutically acceptablesalts thereof;

[0054] and further wherein the compound of Formula (I) has a bindingaffinity to VLA-4 as expressed by an IC₅₀ of about 15 μM or less.

[0055] In a preferred embodiment, Alk is preferably 1 carbon and m ispreferably 1.

[0056] In a preferred embodiment, HetAr in the above compounds is anitrogen containing substituted heteroaryl group, preferably a nitrogencontaining heteroaryl group that is substituted with a substituentselected from the group consisting of acyl, acylamino, acyloxy,aminoacyl, aminocarbonylamino, aminothiocarbonylamino, aminocarbonyloxy,oxycarbonylamino, oxythiocarbonylamino, thioamidino, thiocarbonylamino,aminosulfonylamino, aminosulfonyloxy, aminosulfonyl, oxysulfonylaminooxysulfonyl, aryl and substituted aryl.

[0057] Within this group a more preferred group is wherein the nitrogencontaining heteroaryl group is substituted with a substituent of formula—O-Z-NR¹¹R^(11′) or —O-Z-R¹² wherein R¹¹ and R^(11′) are independentlyselected from the group consisting of hydrogen, alkyl, substitutedalkyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substitutedcycloalkenyl, heterocyclic, substituted heterocyclic, and where R¹¹ andR^(11′) are joined to form a heterocycle or a substituted heterocycle,R¹² is selected from the group consisting of heterocycle and substitutedheterocycle, and Z is selected from the group consisting of —C(O)— and—SO₂—. More preferably, the nitrogen containing heteroaryl group issubstituted with a group of formula —OC(O)NR¹¹R^(11′), wherein R¹¹ andR^(11′) are as defined herein. Even more preferably —OC(O)NR¹¹R^(11′)wherein R¹¹ and R^(11′) are independently selected from the groupconsisting of alkyl or R¹¹ and R^(11′) are joined to form a heterocycleor a substituted heterocycle, most preferably —OC(O)N(CH₃)₂.

[0058] Another more preferred group is wherein the nitrogen containingheteroaryl group is substituted with an aryl or substituted arylsubstituent. More preferably, the heteroaryl group is substituted withdialkoxyphenyl.

[0059] In yet another preferred embodiment, A in the above compounds isheteroaryl optionally substituted with 1 to 3 substituents selected fromthe group consisting of alkyl, substituted alkyl, alkoxy, substitutedalkoxy, amino, substituted amino, cycloalkyl, substituted cycloalkyl,aryl, substituted aryl, heteroaryl, substituted heteroaryl,heterocyclic, substituted heterocyclic and halogen. Preferably, A isselected from the group consisting of 1-oxo-1,2,5-thiadiazole,1,1-dioxo-1,2,5-thiadiazole, pyridazine, pyrimidine or pyrazine; morepreferably, pyrimidine or pyrazine; wherein the pyridazine, pyrimidineor pyrazine ring is optionally substituted with 1 to 3 substituentsselected from the group consisting of alkyl, substituted alkyl, alkoxy,substituted alkoxy, amino, substituted amino, cycloalkyl, substitutedcycloalkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl,heterocyclic, substituted heterocyclic and halogen.

[0060] In yet another preferred embodiment, R¹ is hydrogen, and X ishydroxyl.

[0061] In still another preferred embodiment, this invention is directedto compounds of Formula IIa, IIb, IIc, IId, or IIe:

[0062] wherein:

[0063] HetAr is a nitrogen containing heteroaryl group substituted atany position capable of substitution with a single substitution selectedfrom the group consisting of acyl, acylamino, acyloxy, aminoacyl,aminocarbonylamino, aminothiocarbonylamino, aminocarbonyloxy,oxycarbonylamino, oxythiocarbonylamino, thioamidino, thiocarbonylamino,aminosulfonylamino, aminosulfonyloxy, aminosulfonyl, oxysulfonylamino,aryl, substituted aryl, and oxysulfonyl;

[0064] R⁵ is selected from the group consisting of alkyl, substitutedalkyl, alkenyl, substituted alkenyl, aryl, substituted aryl, cycloalkyl,substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl,heterocyclic, substituted heterocylic, heteroaryl and substitutedheteroaryl;

[0065] R⁶ is selected from the group consisting of hydrogen, alkyl,substituted alkyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl,substituted cycloalkenyl, heterocyclic, substituted heterocyclic, aryl,substituted aryl, heteroaryl, substituted heteroaryl, and —SO₂R¹⁰ whereR¹⁰ is selected from the group consisting of alkyl, substituted alkyl,cycloalkyl, substituted cycloalkyl, cycloalkenyl, substitutedcycloalkenyl, heterocyclic, substituted heterocyclic, aryl, substitutedaryl, heteroaryl, substituted heteroaryl;

[0066] R⁷ and R⁸ are independently selected from the group consisting ofhydrogen, alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl,aryl, substituted aryl, heteroaryl, substituted heteroaryl,heterocyclic, substituted heterocyclic and halogen;

[0067] R¹⁶ and R¹⁷ are independently selected from the group consistingof hydrogen, alkyl, substituted alkyl, alkoxy, substituted alkoxy,amino, substituted amino, cycloalkyl, substituted cycloalkyl, aryl,substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic,substituted heterocyclic and halogen; and

[0068] R¹⁸ is selected from the group consisting of alkyl, substitutedalkyl, alkoxy, substituted alkoxy, amino, substituted amino, cycloalkyl,substituted cycloalkyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic;

[0069] R²⁰ is selected from the group consisting of hydrogen, alkyl,substituted alkyl, alkoxy, substituted alkoxy, cycloalkyl, substitutedcycloalkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl,heterocyclic, substituted heterocyclic and halogen;

[0070] R²¹ is selected from the group consisting of alkyl, substitutedalkyl, alkoxy, substituted alkoxy, amino, substituted amino, cycloalkyl,substituted cycloalkyl, aryl, substituted aryl, heterocyclic andsubstituted heterocyclic;

[0071] b is 1 or 2; and

[0072] X is hydroxyl; and

[0073] and enantiomers, diastereomers and pharmaceutically acceptablesalts thereof.

[0074] Preferably, the compound is selected from Formula IIc, IId orIIe.

[0075] In the above compounds II(a-e), HetAr is preferably:

[0076] a) a nitrogen containing heteroaryl ring which is substitutedwith a group of formula —O-Z-NR¹¹R^(11′) or —O-Z-R¹² wherein R¹¹ andR^(11 ′) are independently selected from the group consisting ofhydrogen, alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl,cycloalkenyl, substituted cycloalkenyl, heterocyclic, substitutedheterocyclic, and where R¹¹ and R^(11′) are joined to form a heterocycleor a substituted heterocycle, R¹² is selected from the group consistingof heterocycle and substituted heterocycle, and Z is selected from thegroup consisting of —C(O)— and —SO₂—. More preferably, the nitrogencontaining heteroaryl ring is substituted with a group of formula—OC(O)NR¹¹R^(11′), wherein R¹¹ and R^(11′) are as defined herein. Evenmore preferably —OC(O)NR¹¹R^(11′) wherein R¹¹ and R^(11′) areindependently selected from the group consisting of alkyl or R¹¹ andR^(11′) are joined to form a heterocycle or a substituted heterocycle,most preferably —OC(O)N(CH₃)₂; or

[0077] b) a nitrogen containing heteroaryl ring which is substitutedwith an aryl or substituted aryl group. More preferably, wherein theheteroaryl ring is substituted with dialkoxyphenyl.

[0078] Another preferred group of compounds are those wherein R⁵ isselected from the group consisting of alkyl, substituted alkyl, aryl,substituted aryl, heterocyclic, substituted heterocylic, heteroaryl andsubstituted heteroaryl. Even more preferably R⁵ is selected from thegroup consisting of 4-methylphenyl, methyl, benzyl, n-butyl, n-hexyl,4-chlorophenyl, 1-naphthyl, 2-naphthyl, 4-methoxyphenyl, phenyl,2,4,6-trimethylphenyl, 2-(methoxycarbonyl)phenyl, 2-carboxyphenyl,3,5-dichlorophenyl, 4-trifluoromethylphenyl, 3,4-dichlorophenyl,3,4-dimethoxyphenyl, 4-(CH₃C(O)NH—)phenyl, 4-trifluoromethoxyphenyl,4-cyanophenyl, isopropyl, 3,5-di-(trifluoromethyl)phenyl,4-t-butylphenyl, 4-t-butoxyphenyl, 4-nitrophenyl, 2-thienyl,1-N-methyl-3-methyl-5-chloropyrazol-4-yl, phenethyl,1-N-methylimidazol-4-yl, 4-bromophenyl, 4-amidinophenyl,4-methylamidinophenyl, 4-[CH₃SC(═NH)]phenyl, 5-chloro-2-thienyl,2,5-dichloro-4-thienyl, 1-N-methyl-4-pyrazolyl, 2-thiazolyl,5-methyl-1,3,4-thiadiazol-2-yl, 4-[H₂NC(S)]phenyl, 4-aminophenyl,4-fluorophenyl, 2-fluorophenyl, 3-fluorophenyl, 3,5-difluorophenyl,pyridin-3-yl, pyrimidin-2-yl, 4-(3′-dimethylamino-n-propoxy)-phenyl, and1-methylpyrazol-4-yl;

[0079] R¹⁶ is substituted amino;

[0080] R⁶, R¹⁷ and/or R²⁰ are hydrogen; and

[0081] R¹⁸ and/or R²¹ are alkyl, substituted alkyl, aryl, or substitutedaryl.

[0082] In a second aspect, this invention provides pharmaceuticalcompositions comprising a pharmaceutically acceptable carrier and atherapeutically effective amount of the compounds defined herein.

[0083] In a third aspect, this invention is directed to a method fortreating a disease mediated by VLA-4 in a patient, which methodcomprises administering a pharmaceutical composition comprising apharmaceutically acceptable carrier and a therapeutically effectiveamount of compounds defined herein.

[0084] The compounds and pharmaceutical compositions of this inventionare useful for treating disease conditions mediated by VLA-4 orleucocyte adhesion. Such disease conditions include, by way of example,asthma, Alzheimer's disease, atherosclerosis, AIDS dementia, diabetes(including acute juvenile onset diabetes), inflammatory bowel disease(including ulcerative colitis and Crohn's disease), multiple sclerosis,rheumatoid arthritis, tissue transplantation, tumor metastasis,meningitis, encephalitis, stroke, and other cerebral traumas, nephritis,retinitis, atopic dermatitis, psoriasis, myocardial ischemia and acuteleukocyte-mediated lung injury such as that which occurs in adultrespiratory distress syndrome.

[0085] Other disease conditions include, but are not limited to,inflammatory conditions such as erythema nodosum, allergicconjunctivitis, optic neuritis, uveitis, allergic rhinitis, Ankylosingspondylitis, psoriatic arthritis, vasculitis, Reiter's syndrome,systemic lupus erythematosus, progressive systemic sclerosis,polymyositis, dermatomyositis, Wegner's granulomatosis, aortitis,sarcoidosis, lymphocytopenia, temporal arteritis, pericarditis,myocarditis, congestive heart failure, polyarteritis nodosa,hypersensitivity syndromes, allergy, hypereosinophilic syndromes,Churg-Strauss syndrome, chronic obstructive pulmonary disease,hypersensitivity pneumonitis, chronic active hepatitis, interstitialcystitis, autoimmune endocrine failure, primary biliary cirrhosis,autoimmune aplastic anemia, chronic persistent hepatitis andthyroiditis.

[0086] In a preferred embodiment, the disease condition mediated byVLA-4 is an inflammatory disease.

[0087] In the above compounds, when X is other than —OH orpharmaceutical salts thereof, X is preferably a substituent which willconvert (e.g., hydrolyze, metabolize, etc.) in vivo to a compound whereX is —OH or a salt thereof. Accordingly, suitable X groups are any artrecognized pharmaceutically acceptable groups which will hydrolyze orotherwise convert in vivo to a hydroxyl group or a salt thereofincluding, by way of example, esters (X is alkoxy, substituted alkoxy,cycloalkoxy, substituted cycloalkoxy, alkenoxy, substituted alkenoxy,cycloalkenoxy, substituted cycloalkenoxy, aryloxy, substituted aryloxy,heteroaryloxy, substituted heteroaryloxy, heterocyclooxy, substitutedheterocyclooxy, and the like).

[0088] This invention also provides methods for binding VLA-4 in abiological sample which method comprises contacting the biologicalsample with a compound of this invention under conditions wherein saidcompound binds to VLA-4.

[0089] The pharmaceutical compositions may be used to treat diseaseconditions mediated by VLA-4 or leucocyte adhesion. Such diseaseconditions include, by way of example, asthma, Alzheimer's disease,atherosclerosis, AIDS dementia, diabetes (including acute juvenile onsetdiabetes), inflammatory bowel disease (including ulcerative colitis andCrohn's disease), multiple sclerosis, rheumatoid arthritis, tissuetransplantation, tumor metastasis, meningitis, encephalitis, stroke, andother cerebral traumas, nephritis, retinitis, atopic dermatitis,psoriasis, myocardial ischemia and acute leukocyte-mediated lung injurysuch as that which occurs in adult respiratory distress syndrome.

[0090] Other disease conditions include, but are not limited to,inflammatory conditions such as erythema nodosum, allergicconjunctivitis, optic neuritis, uveitis, allergic rhinitis, ankylosingspondylitis, psoriatic arthritis, vasculitis, Reiter's syndrome,systemic lupus erythematosus, progressive systemic sclerosis,polymyositis, dermatomyositis, Wegner's granulomatosis, aortitis,sarcoidosis, lymphocytopenia, temporal arteritis, pericarditis,myocarditis, congestive heart failure, polyarteritis nodosa,hypersensitivity syndromes, allergy, hypereosinophilic syndromes,Churg-Strauss syndrome, chronic obstructive pulmonary disease,hypersensitivity pneumonitis, chronic active hepatitis, interstitialcystitis, autoimmune endocrine failure, primary biliary cirrhosis,autoimmune aplastic anemia, chronic persistent hepatitis andthyroiditis.

[0091] Preferred compounds of this invention include those set forth inTable I below: TABLE I

Cpd # A HetAr 1 [2-N(CH₃)₂-5-CF₃CH₂]pyrimidin-4-yl2-(2,6-di-MeOPh)pyridin-5-yl 2 [2-N(CH₃)₂-5-CF₃CH₂]pyrimidin-4-yl5-(2,6-di-MeOPh)pyridin-2-yl 3 [2-N(CH₃)₂-5-CF₃CH₂]pyrimidin-4-yl3-(2,6-di-MeOPh)pyridazin-6-yl 4 [2-N(CH₃)₂-5-CF₃CH₂]pyrimidin-4-yl5-(2,6-di-MeOPh)pyrimidin-5-yl 5 [2-N(CH₃)₂-5-CF₃CH₂]pyrimidin-4-yl2-(2,6-di-MeOPh)pyrimidin-5-yl 6 [2-N(CH₃)₂-5-CF₃CH₂]pyrimidin-4-yl5-(2,6-di-MeOPh)pyrazin-2-yl 7 [2-N(CH₃)₂-5-(CH₃)₂CH]pyrimidin-4-yl2-(2,6-di-MeOPh)pyridin-5-yl 8 [2-N(CH₃)₂-5-(CH₃)₂CH]pyrimidin-4-yl5-(2,6-di-MeOPh)pyridin-2-yl 9 [2-N(CH₃)₂-5-(CH₃)₂CH]pyrimidin-4-yl2-(2,6-di-MeOPh)pyridazin-6-yl 10 [2-N(CH₃)₂-5-(CH₃)₂CH]pyrimidin-4-yl5-(2,6-di-MeOPh)pyrimidin-2-yl 11 [2-N(CH₃)₂-5-(CH₃)₂CH]pyrimidin-4-yl2-(2,6-di-MeOPh)pyrimidin-5-yl 12 [2-N(CH₃)₂-5-(CH₃)₂CH]pyrimidin-4-yl5-(2,6-di-MeOPh)pyrazin-2-yl 13 [2-N(CH₃)₂-5-(CH₃CH₂)₂CH]-pyrimidin-4-yl2-(2,6-di-MeOPh)pyridin-5-yl 14 [2-N(CH₃)₂-5-(CH₃CH₂)₂CH]-pyrimidin-4-yl5-(2,6-di-MeOPh)pyridin-2-yl 15 [2-N(CH₃)₂-5-(CH₃CH₂)₂CH]-pyrimidin-4-yl3-(2,6-di-MeOPh)pyridazin-6-yl 16[2-N(CH₃)₂-5-(CH₃CH₂)₂CH]-pyrimidin-4-yl 2-(2,6-di-MeOPh)pyrimidin-2-yl17 [2-N(CH₃)₂-5-(CH₃CH₂)₂CH]-pyrimidin-4-yl2-(2,6-di-MeOPh)pyrimidin-5-yl 18[2-N(CH₃)₂-5-(CH₃CH₂)₂CH]-pyrimidin-4-yl 5-(2,6-di-MeOPh)pyriazin-2-yl19 [2-N(CH₃)₂-5-(3,5-di-CH₃isoxazol-4-yl]pyrimidin-4-yl2-(2,6-di-MeOPh)pyridin-5-yl 20[2-N(CH₃)₂-5-(3,5-di-CH₃isoxazol-4-yl]pyrimidin-4-yl5-(2,6-di-MeOPh)pyridin-2-yl 21[2-N(CH₃)₂-5-(3,5-di-CH₃isoxazol-4-yl]pyrimidin-4-yl3-(2,6-di-MeOPh)pyridazin-6-yl 22[2-N(CH₃)₂-5-(3,5-di-CH₃isoxazol-4-yl]pyrimidin-4-yl5-(2,6-di-MeOPh)pyrimidin-2-yl 23[2-N(CH₃)₂-5-(3,5-di-CH₃isoxazol-4-yl]pyrimidin-4-yl2-(2,6-di-MeOPh)pyrimidin-5-yl 24[2-N(CH₃)₂-5-(3,5-di-CH₃isoxazol-4-yl]pyrimidin-4-yl5-(2,6-di-MeOPh)pyrazin-2-yl 25[2-N(CH₃)₂-5-(1,3,5-tri-CH₃pyrazol-4-yl]pyrimidin-4-yl2-(2,6-di-MeOPh)pyridin-5-yl 26[2-N(CH₃)₂-5-(1,3,5-tri-CH₃pyrazol-4-yl]pyrimidin-4-yl5-(2,6-di-MeOPh)pyridin-2-yl 27[2-N(CH₃)₂-5-(1,3,5-tri-CH₃pyrazol-4-yl]pyrimidin-4-yl3-(2,6-di-MeOPh)pyridazin-6-yl 28[2-N(CH₃)₂-5-(1,3,5-tri-CH₃pyrazol-4-yl]pyrimidin-4-yl5-(2,6-di-MeOPh)pyrimidin-2-yl 29[2-N(CH₃)₂-5-(1,3,5-tri-CH₃pyrazol-4-yl]pyrimidin-4-yl2-(2,6-di-MeOPh)pyrimidin-5-yl 30[2-N(CH₃)₂-5-(1,3,5-tri-CH₃pyrazol-4-yl]pyrimidin-4-yl5-(2,6-di-MeOPh)pyrazin-2-yl 31[2-N(CH₃)₂-5-(3,5-di-CH₃isothiazol-4-yl]pyrimidin-4-yl2-(2,6-di-MeOPh)pyridin-5-yl 32[2-N(CH₃)₂-5-(3,5-di-CH₃isothiazol-4-yl]pyrimidin-4-yl5-(2,6-di-MeOPh)pyridin-2-yl 33[2-N(CH₃)₂-5-(3,5-di-CH₃isothiazol-4-yl]pyrimidin-4-yl3-(2,6-di-MeOPh)pyridazin-6-yl 34[2-N(CH₃)₂-5-(3,5-di-CH₃isothiazol-4-yl]pyrimidin-4-yl5-(2,6-di-MeOPh)pyrimidin-2-yl 35[2-N(CH₃)₂-5-(3,5-di-CH₃isothiazol-4-yl]pyrimidin-4-yl2-(2,6-di-MeOPh)pyrimidin-5-yl 36[2-N(CH₃)₂-5-(3,5-di-CH₃isothiazol-4-yl]pyrimidin-4-yl5-(2,6-di-MeOPh)pyriazin-2-yl

[0092] and are named as follows:

[0093] N-(2-(N,N-dimethylamino)-5-(2,2,2-trifluoroethyl)pyrimidin-4-yl)-L-3-(2-(2,6-dimethoxyphenyl)pyridin-5-yl)alanine;

[0094]N-(2-(N,N-dimethylamino)-5-(2,2,2-trifluoroethyl)pyrimidin-4-yl)-L-3-(5-(2,6-dimethoxyphenyl)pyridin-2-yl)alanine;

[0095]N-(2-(N,N-dimethylamino)-5-(2,2,2-trifluoroethyl)pyrimidin-4-yl)-L-3-(3-(2,6-dimethoxyphenyl)pyridazin-6-yl)alanine;

[0096]N-(2-(N,N-dimethylamino)-5-(2,2,2-trifluoroethyl)pyrimidin-4-yl)-L-3-(5-(2,6-dimethoxyphenyl)pyrimidin-2-yl)alanine;

[0097]N-(2-(N,N-dimethylamino)-5-(2,2,2-trifluoroethyl)pyrimidin-4-yl)-L-3-(2-(2,6-dimethoxyphenyl)pyridin-5-yl)alanine;

[0098]N-(2-(N,N-dimethylamino)-5-(2-methylpropyl)pyrimidin-4-yl)-L-3-(2-(2,6-dimethoxyphenyl)pyrimidin-5-yl)alanine;

[0099]N-(2-(N,N-dimethylamino)-5-(2,2,2-trifluoroethyl)pyrimidin-4-yl)-L-3-(5-(2,6-dimethoxyphenyl)pyrazin-2-yl)alanine;

[0100]N-(2-(N,N-dimethylamino)-5-(2-methylpropyl)pyrimidin-4-yl)-L-3-(2-(2,6-dimethoxyphenyl)pyridin-5-yl)alanine;

[0101]N-(2-(N,N-dimethylamino)-5-(2-methypropyl)pyrimidin-4-yl)-L-3-(5-(2,6-dimethoxyphenyl)pyridin-2-yl)alanine;

[0102]N-(2-(N,N-dimethylamino)-5-(2-methylpropyl)pyrimidin-4-yl)-L-3-(3-(2,6-dimethoxyphenyl)pyridazin-6-yl)alanine;

[0103] N-(2-(N,N-dimethylamino)-5-(2-methylpropyl)pyrimidin-4-yl)-L-3-(5-(2,6-dimethoxyphenyl)pyrimidin-2-yl)alanine;

[0104]N-(2-(N,N-dimethylamino)-5-(2-methylpropyl)pyrimidin-4-yl)-L-3-(2-(2,6-dimethoxyphenyl)pyrimidin-5-yl)alanine;

[0105]N-(2-(N,N-dimethylamino)-5-(2-methylpropyl)pyrimidin-4-yl)-L-3-(52-(2,6-dimethoxyphenyl)pyrazin-2-yl)alanine;

[0106]N-(2-(N,N-dimethylamino)-5-(1-ethylpropyl)pyrimidin-4-yl)-L-3-(2-(2,6-dimethoxyphenyl)pyridin-5-yl)alanine;

[0107]N-(2-(N,N-dimethylamino)-5-(1-ethylpropyl)pyrimidin-4-yl)-L-3-(5-(2,6-dimethoxyphenyl)pyridin-2-yl)alanine;

[0108]N-(2-(N,N-dimethylamino)-5-(1-ethylpropyl)pyrimidin-4-yl)-L-3-(3-(2,6-dimethoxyphenyl)pyridazin-6-yl)alanine;

[0109]N-(2-(N,N-dimethylamino)-5-(1-ethylpropyl)pyrimidin-4-yl)-L-3-(5-(2,6-dimethoxyphenyl)pyrimidin-2-yl)alanine;

[0110]N-(2-(N,N-dimethylamino)-5-(1-ethylpropyl)pyrimidin-4-yl)-L-3-(2-(2,6-dimethoxyphenyl)pyrimidin-5-yl)alanine;

[0111]N-(2-(N,N-dimethylamino)-5-(1-ethylpropyl)pyrimidin-4-yl)-L-3-(5-(2,6-dimethoxyphenyl)pyrazin-2-yl)alanine;

[0112]N-(2-(N,N-dimethylamino)-5-(3,5-dimethylisoxazol-4-yl)pyrimidin-4-yl)-L-3-(2-(2,6-dimethoxyphenyl)pyridin-5-yl)alanine;

[0113]N-(2-(N,N-dimethylamino)-5-(3,5-dimethylisoxazol-4-yl)pyrimidin-4-yl)-L-3-(5-(2,6-dimethoxyphenyl)pyridin-2-yl)alanine;

[0114]N-(2-(N,N-dimethylamino)-5-(3,5-dimethylisoxazol-4-yl)pyrimidin-4-yl)-L-3-(3-(2,6-dimethoxyphenyl)pyridazin-6-yl)alanine;

[0115]N-(2-(N,N-dimethylamino)-5-(3,5-dimethylisoxazol-4-yl)pyrimidin-4-yl)-L-3-(5-(2,6-dimethoxyphenyl)pyrimidin-2-yl)alanine;

[0116]N-(2-(N,N-dimethylamino)-5-(3,5-dimethylisoxazol-4-yl)pyrimidin-4-yl)-L-3-(2-(2,6-dimethoxyphenyl)pyrimidin-5-yl)alanine;

[0117]N-(2-(N,N-dimethylamino)-5-(3,5-dimethylisoxazol-4-yl)pyrimidin-4-yl)-L-3-(5-(2,6-dimethoxyphenyl)pyrazin-2-yl)alanine;

[0118]N-(2-(N,N-dimethylamino)-5-(1,3,5-trimethylpyrazol-4-yl)pyrimidin-4-yl)-L-3-(2-(2,6-dimethoxyphenyl)pyridin-5-yl)alanine;

[0119]N-(2-(N,N-dimethylamino)-5-(1,3,5-trimethylpyrazol-4-yl)pyrimidin-4-yl)-L-3-(5-(2,6-dimethoxyphenyl)pyridin-2-yl)alanine;

[0120]N-(2-(N,N-dimethylamino)-5-(1,3,5-trimethylpyrazol-4-yl)pyrimidin-4-yl)-L-3-(3-(2,6-dimethoxyphenyl)pyridazin-6-yl)alanine;

[0121]N-(2-(N,N-dimethylamino)-5-(1,3,5-trimethylpyrazol-4-yl)pyrimidin-4-yl)-L-3-(5-(2,6-dimethoxyphenyl)pyrimidin-2-yl)alanine;

[0122]N-(2-(N,N-dimethylamino)-5-(1,3,5-trimethylpyrazol-4-yl)pyrimidin-4-yl)-L-3-(2-(2,6-dimethoxyphenyl)pyrimidin-5-yl)alanine;

[0123]N-(2-(N,N-dimethylamino)-5-(1,3,5-trimethylpyrazol-4-yl)pyrimidin-4-yl)-L-3-(5-(2,6-dimethoxyphenyl)pyrazin-2-yl)alanine;

[0124]N-(2-(N,N-dimethylamino)-5-(3,5-dimethylisothiazol-4-yl)pyrimidin-4-yl)-L-3-(2-(2,6-dimethoxyphenyl)pyridin-5-yl)alanine;

[0125]N-(2-(N,N-dimethylamino)-5-(1,3,5-trimethylpyrazol-4-yl)pyrimidin-4-yl)-L-3-(5-(2,6-dimethoxyphenyl)pyridin-2-yl)alanine;

[0126]N-(2-(N,N-dimethylamino)-5-(1,3,5-trimethylpyrazol-4-yl)pyrimidin-4-yl)-L-3-(3-(2,6-dimethoxyphenyl)pyridazin-6-yl)alanine;

[0127]N-(2-(N,N-dimethylamino)-5-(1,3,5-trimethylpyrazol-4-yl)pyrimidin-4-yl)-L-3-(5-(2,6-dimethoxyphenyl)pyrimidin-2-yl)alanine;

[0128]N-(2-(N,N-dimethylamino)-5-(1,3,5-trimethylpyrazol-4-yl)pyrimidin-4-yl)-L-3-(2-(2,6-dimethoxyphenyl)pyrimidin-5-yl)alanine;and

[0129]N-(2-(N,N-dimethylamino)-5-(1,3,5-trimethylpyrazol-4-yl)pyrimidin-4-yl)-L-3-(5-(2,6-dimethoxyphenyl)pyrazin-2-yl)alanine;

[0130] including all pharmaceutically acceptable salts thereof.

DETAILED DESCRIPTION OF THE INVENTION

[0131] As above, this invention relates to compounds which inhibitleukocyte adhesion and, in particular, leukocyte adhesion mediated byVLA-4. However, prior to describing this invention in further detail,the following terms will first be defined.

[0132] Definitions

[0133] As used herein, “alkyl” refers to alkyl groups preferably havingfrom 1 to 10 carbon atoms and more preferably 1 to 6 carbon atoms. Thisterm is exemplified by groups such as methyl, t-butyl, n-heptyl, octyland the like.

[0134] “Alkylene” refers to alkylene groups preferably having from 1 to4 carbon atoms.

[0135] “Substituted alkyl” refers to an alkyl group, preferably of from1 to 10 carbon atoms, having from 1 to 5 substituents selected from thegroup consisting of alkoxy, substituted alkoxy, acyl, acylamino,thiocarbonylamino, acyloxy, amino, amidino, alkyl amidino,thioamidino,aminoacyl, aminocarbonylamino, aminothiocarbonylamino, aminocarbonyloxy,aryl, substituted aryl, aryloxy, substituted aryloxy, aryloxyaryl,substituted aryloxyaryl, cyano, halogen, hydroxyl, nitro, carboxyl,carboxylalkyl, carboxyl-substituted alkyl, carboxyl-cycloalkyl,carboxyl-substituted cycloalkyl, carboxylaryl, carboxyl-substitutedaryl, carboxylheteroaryl, carboxyl-substituted heteroaryl,carboxylheterocyclic, carboxyl-substituted heterocyclic, cycloalkyl,substituted cycloalkyl, guanidino, guanidinosulfone, thiol, thioalkyl,substituted thioalkyl, thioaryl, substituted thioaryl, thiocycloalkyl,substituted thiocycloalkyl, thioheteroaryl, substituted thioheteroaryl,thioheterocyclic, substituted thioheterocyclic, heteroaryl, substitutedheteroaryl, heterocyclic, substituted heterocyclic, cycloalkoxy,substituted cycloalkoxy, heteroaryloxy, substituted heteroaryloxy,heterocyclyloxy, substituted heterocyclyloxy, oxycarbonylamino,oxythiocarbonylamino, —OS(O)₂-alkyl, —OS(O)₂-substituted alkyl,—OS(O)₂-aryl, —OS(O)₂-substituted aryl, —OS(O)₂-heteroaryl,—OS(O)₂-substituted heteroaryl, —OS(O)₂-heterocyclic,—OS(O)₂-substituted heterocyclic, —OSO₂—NRR where R is hydrogen oralkyl, —NRS(O)₂-alkyl, —NRS(O)₂-substituted alkyl, —NRS(O)₂-aryl,—NRS(O)₂-substituted aryl, —NRS(O)₂-heteroaryl, —NRS(O)₂-substitutedheteroaryl, —NRS(O)₂-heterocyclic, —NRS(O)₂-substituted heterocyclic,—NRS(O)₂—NR-alkyl, —NRS(O)₂—NR-substituted alkyl, —NRS(O)₂—NR-aryl,—NRS(O)₂—NR-substituted aryl, —NRS(O)₂—NR-heteroaryl,—NRS(O)₂—NR-substituted heteroaryl, —NRS(O)₂—NR-heterocyclic,—NRS(O)₂—NR-substituted heterocyclic where R is hydrogen or alkyl, mono-and di-alkylamino, mono- and di-(substituted alkyl)amino, mono- anddi-arylamino, mono- and di-substituted arylamino, mono- anddi-heteroarylamino, mono- and di-substituted heteroarylamino, mono- anddi-heterocyclic amino, mono- and di-substituted heterocyclic amino,unsymmetric di-substituted amines having different substituents selectedfrom alkyl, substituted alkyl, aryl, substituted aryl, heteroaryl,substituted heteroaryl, heterocyclic and substituted heterocyclic andsubstituted alkyl groups having amino groups blocked by conventionalblocking groups such as Boc, Cbz, formyl, and the like oralkyl/substituted alkyl groups substituted with —SO₂-alkyl,—SO₂-substituted alkyl, —SO₂-alkenyl, —SO₂-substituted alkenyl,—SO₂-cycloalkyl, —SO₂-substituted cycloalkyl, —SO₂-aryl,—SO₂-substituted aryl, —SO₂-heteroaryl, —SO₂-substituted heteroaryl,—SO₂-heterocyclic, —SO₂-substituted heterocyclic and —SO₂NRR where R ishydrogen or alkyl.

[0136] “Alkoxy” refers to the group “alkyl-O—” which includes, by way ofexample, methoxy, ethoxy, n-propoxy, iso-propoxy, n-butoxy, tert-butoxy,sec-butoxy, n-pentoxy, n-hexoxy, 1,2-dimethylbutoxy, and the like.

[0137] “Substituted alkoxy” refers to the group “substituted alkyl-O—”.

[0138] “Alkenoxy” refers to the group “alkenyl-O—”.

[0139] “Substituted alkenoxy” refers to the group “substitutedalkenyl-O—”.

[0140] “Acyl” refers to the groups H—C(O)—, alkyl-C(O)—, substitutedalkyl-C(O)—, alkenyl-C(O)—, substituted alkenyl-C(O)—, alkynyl-C(O)—,substituted alkynyl-C(O)— cycloalkyl-C(O)—, substitutedcycloalkyl-C(O)—, aryl-C(O)—, substituted aryl-C(O)—, heteroaryl-C(O)—,substituted heteroaryl-C(O), heterocyclic-C(O)—, and substitutedheterocyclic-C(O)— wherein alkyl, substituted alkyl, alkenyl,substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl,substituted cycloalkyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

[0141] “Acylamino” refers to the group —C(O)NRR where each R isindependently selected from the group consisting of hydrogen, alkyl,substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substitutedalkynyl, aryl, substituted aryl, cycloalkyl, substituted cycloalkyl,heteroaryl, substituted heteroaryl, heterocyclic, substitutedheterocyclic and where each R is joined to form together with thenitrogen atom a heterocyclic or substituted heterocyclic ring whereinalkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl,substituted alkynyl, cycloalkyl, substituted cycloalkyl, aryl,substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic andsubstituted heterocyclic are as defined herein.

[0142] “Thiocarbonylamino” refers to the group —C(S)NRR where each R isindependently selected from the group consisting of hydrogen, alkyl,substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substitutedalkynyl, aryl, substituted aryl, cycloalkyl, substituted cycloalkyl,heteroaryl, substituted heteroaryl, heterocyclic, substitutedheterocyclic and where each R is joined to form, together with thenitrogen atom a heterocyclic or substituted heterocyclic ring whereinalkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl,substituted alkynyl, cycloalkyl, substituted cycloalkyl, aryl,substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic andsubstituted heterocyclic are as defined herein.

[0143] “Acyloxy” refers to the groups alkyl-C(O)O—, substitutedalkyl-C(O)O—, alkenyl-C(O)O—, substituted alkenyl-C(O)O—,alkynyl-C(O)O—, substituted alkynyl-C(O)O—, aryl-C(O)O—, substitutedaryl-C(O)O—, cycloalkyl-C(O)O—, substituted cycloalkyl-C(O)O—,heteroaryl-C(O)O—, substituted heteroaryl-C(O)O—, heterocyclic-C(O)O—,and substituted heterocyclic-C(O)O— wherein alkyl, substituted alkyl,alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl,substituted cycloalkyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

[0144] “Oxysulfonyl” refers to the groups alkyl-SO₂O—, substitutedalkyl-SO₂O—, alkenyl-SO₂O—, substituted alkenyl-SO₂O—, alkynyl-SO₂O—,substituted alkynyl-SO₂O—, aryl-SO₂O—, substituted aryl-SO₂O—,cycloalkyl-SO₂O—, substituted cycloalkyl-SO₂O—, heteroaryl-SO₂O—,substituted heteroaryl-SO₂O—, heterocyclic-SO₂O—, and substitutedheterocyclic-SO₂O— wherein alkyl, substituted alkyl, alkenyl,substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl,substituted cycloalkyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

[0145] “Alkenyl” refers to alkenyl group preferably having from 2 to 10carbon atoms and more preferably 2 to 6 carbon atoms and having at least1 and preferably from 1-2 sites of alkenyl unsaturation.

[0146] “Substituted alkenyl” refers to alkenyl groups having from 1 to 5substituents selected from the group consisting of alkoxy, substitutedalkoxy, acyl, acylamino, thiocarbonylamino, acyloxy, amino, amidino,alkylamidino, thioamidino, aminoacyl, aminocarbonylamino,aminothiocarbonylamino, aminocarbonyloxy, aryl, substituted aryl,aryloxy, substituted aryloxy, aryloxyaryl, substituted aryloxyaryl,halogen, hydroxyl, cyano, nitro, carboxyl, carboxylalkyl,carboxyl-substituted alkyl, carboxyl-cycloalkyl, carboxyl-substitutedcycloalkyl, carboxylaryl, carboxyl-substituted aryl, carboxylheteroaryl,carboxyl-substituted heteroaryl, carboxylheterocyclic,carboxyl-substituted heterocyclic, cycloalkyl, substituted cycloalkyl,guanidino, guanidinosulfone, thiol, thioalkyl, substituted thioalkyl,thioaryl, substituted thioaryl, thiocycloalkyl, substitutedthiocycloalkyl, thioheteroaryl, substituted thioheteroaryl,thioheterocyclic, substituted thioheterocyclic, heteroaryl, substitutedheteroaryl, heterocyclic, substituted heterocyclic, cycloalkoxy,substituted cycloalkoxy, heteroaryloxy, substituted heteroaryloxy,heterocyclyloxy, substituted heterocyclyloxy, oxycarbonylamino,oxythiocarbonylamino, —OS(O)₂-alkyl, —OS(O)₂-substituted alkyl,—OS(O)₂-aryl, —OS(O)₂-substituted aryl, —OS(O)₂-heteroaryl,—OS(O)₂-substituted heteroaryl, —OS(O)₂-heterocyclic,—OS(O)₂-substituted heterocyclic, —OSO₂—NRR where R is hydrogen oralkyl, —N RS(O)₂-alkyl, —NRS(O)₂-substituted alkyl, —NRS(O)₂-aryl,—NRS(O)₂-substituted aryl, —NRS(O)₂-heteroaryl, —NRS(O)₂-substitutedheteroaryl, —NRS(O)₂-heterocyclic, —NRS(O)₂-substituted heterocyclic,—NRS(O)₂—NR-alkyl, —NRS(O)₂-NR-substituted alkyl, —NRS(O)₂—NR-aryl,—NRS(O)₂-NR-substituted aryl, —NRS(O)₂-NR-heteroaryl,—NRS(O)₂—NR-substituted heteroaryl, —NRS(O)₂—NR-heterocyclic,—NRS(O)₂—NR-substituted heterocyclic where R is hydrogen or alkyl, mono-and di-alkylamino, mono- and di-(substituted alkyl)amino, mono- anddi-arylamino, mono- and di-substituted arylamino, mono- anddi-heteroarylamino, mono- and di-substituted heteroarylamino, mono- anddi-heterocyclic amino, mono- and di-substituted heterocyclic amino,unsymmetric di-substituted amines having different substituents selectedfrom alkyl, substituted alkyl, aryl, substituted aryl, heteroaryl,substituted heteroaryl, heterocyclic and substituted heterocyclic andsubstituted alkenyl groups having amrino groups blocked by conventionalblocking groups such as Boc, Cbz, formyl, and the like oralkenyl/substituted alkenyl groups substituted with —SO₂-alkyl,—SO₂-substituted alkyl, —SO₂-alkenyl, —SO₂-substituted alkenyl,—SO₂-cycloalkyl, —SO₂-substituted cycloalkyl, —SO₂-aryl,—SO₂-substituted aryl, —SO₂-heteroaryl, —SO₂-substituted heteroaryl,—SO₂-heterocyclic, —SO₂-substituted heterocyclic and —SO₂NRR where R ishydrogen or alkyl.

[0147] “Alkynyl” refers to alkynyl group preferably having from 2 to 10carbon atoms and more preferably 3 to 6 carbon atoms and having at least1 and preferably from 1-2 sites of alkynyl unsaturation.

[0148] “Substituted alkynyl” refers to alkynyl groups having from 1 to 5substituents selected from the group consisting of alkoxy, substitutedalkoxy, acyl, acylamino, thiocarbonylamino, acyloxy, amino, amidino,alkylamidino, thioamidino, aminoacyl, aminocarbonylamino,aminothiocarbonylamino, aminocarbonyloxy, aryl, substituted aryl,aryloxy, substituted aryloxy, aryloxyaryl, substituted aryloxyaryl,halogen, hydroxyl, cyano, nitro, carboxyl, carboxylalkyl,carboxyl-substituted alkyl, carboxyl-cycloalkyl, carboxyl-substitutedcycloalkyl, carboxylaryl, carboxyl-substituted aryl, carboxylheteroaryl,carboxyl-substituted heteroaryl, carboxylheterocyclic,carboxyl-substituted heterocyclic, cycloalkyl, substituted cycloalkyl,guanidino, guanidinosulfone, thiol, thioalkyl, substituted thioalkyl,thioaryl, substituted thioaryl, thiocycloalkyl, substitutedthiocycloalkyl, thioheteroaryl, substituted thioheteroaryl,thioheterocyclic, substituted thioheterocyclic, heteroaryl, substitutedheteroaryl, heterocyclic, substituted heterocyclic, cycloalkoxy,substituted cycloalkoxy, heteroaryloxy, substituted heteroaryloxy,heterocyclyloxy, substituted heterocyclyloxy, oxycarbonylamino,oxythiocarbonylamino, —OS(O)₂-alkyl, —OS(O)₂-substituted alkyl,—OS(O)₂-aryl, —OS(O)₂-substituted aryl, —OS(O)₂-heteroaryl,—OS(O)₂-substituted heteroaryl, —OS(O)₂-heterocyclic,—OS(O)₂-substituted heterocyclic, —OSO₂—NRR where R is hydrogen oralkyl, —NRS(O)₂-alkyl, —NRS(O)₂-substituted alkyl, —NRS(O)₂-aryl,—NRS(O)₂-substituted aryl, —NRS(O)₂-heteroaryl, —NRS(O)₂-substitutedheteroaryl, —NRS(O)₂-heterocyclic, —NRS(O)₂-substituted heterocyclic,—NRS(O)₂—NR-alkyl, —NRS(O)₂—NR-substituted alkyl, —NRS(O)₂—NR-aryl,—NRS(O)₂—NR-substituted aryl, —NRS(O)₂—NR-heteroaryl,—NRS(O)₂—NR-substituted heteroaryl, —NRS(O)₂—NR-heterocyclic,—NRS(O)₂—NR-substituted heterocyclic where R is hydrogen or alkyl, mono-and di-alkylamino, mono- and di-(substituted alkyl)amino, mono- anddi-arylamino, mono- and di-substituted arylamino, mono- anddi-heteroarylamino, mono- and di-substituted heteroarylamino, mono- anddi-heterocyclic amino, mono- and di-substituted heterocyclic amino,unsymmetric di-substituted amines having different substituents selectedfrom alkyl, substituted alkyl, aryl, substituted aryl, heteroaryl,substituted heteroaryl, heterocyclic and substituted heterocyclic andsubstituted alkynyl groups having amino groups blocked by conventionalblocking groups such as Boc, Cbz, formyl, and the like oralkynyl/substituted alkynyl groups substituted with —SO₂-alkyl,—SO₂-substituted alkyl, —SO₂-alkenyl, —SO₂-substituted alkenyl,—SO₂-cycloalkyl, —SO₂-substituted cycloalkyl, —SO₂-aryl,—SO₂-substituted aryl, —SO₂-heteroaryl, —SO₂-substituted heteroaryl,—SO₂-heterocyclic, —SO₂-substituted heterocyclic and —SO₂NRR where R ishydrogen or alkyl.

[0149] “Amidino” refers to the group H₂NC(═NH)— and the term“alkylamidino” refers to compounds having 1 to 3 alkyl groups (e.g.,alkylHNC(═NH)—).

[0150] “Thioamidino” refers to the group RSC(═NH)— where R is hydrogenor alkyl.

[0151] “Amino” refers to the group —NH₂.

[0152] “Substituted amino” refers to the group —NRR, where each R groupis independently selected from the group consisting of hydrogen, alkyl,substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substitutedalkynyl, cycloalkyl, substituted cycloalkyl, aryl, substituted aryl,heteroaryl, substituted heteroaryl, heterocyclic, substitutedheterocyclic, —SO₂-alkyl, —SO₂-substituted alkyl, —SO₂-alkenyl,—SO₂-substituted alkenyl, —SO₂-cycloalkyl, —SO₂-substituted cycloalkyl,—SO₂-aryl, —SO₂-substituted aryl, —SO₂-heteroaryl, —SO₂-substitutedheteroaryl, —SO₂-heterocyclic, —SO₂-substituted heterocyclic, providedthat both R groups are not hydrogen; or the R groups can be joinedtogether with the nitrogen atom to form a heterocyclic or substitutedheterocyclic ring.

[0153] “Aminoacyl” refers to the groups —NRC(O)alkyl, —NRC(O)substitutedalkyl, —NRC(O)cycloalkyl, —NRC(O)substituted cycloalkyl, —NRC(O)alkenyl,—NRC(O)substituted alkenyl, —NRC(O)alkynyl, —NRC(O)substituted alkynyl,—NRC(O)aryl, —NRC(O)substituted aryl, —NRC(O)heteroaryl,—NRC(O)substituted heteroaryl, —NRC(O)heterocyclic, and—NRC(O)substituted heterocyclic where R is hydrogen or alkyl and whereinalkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl,substituted alkynyl, cycloalkyl, substituted cycloalkyl, aryl,substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic andsubstituted heterocyclic are as defined herein.

[0154] “Aminosulfonyl” refers to the groups —NRSO₂alkyl,—NRSO₂substituted alkyl, —NRSO₂cycloalkyl, —NRSO₂substituted cycloalkyl,—NRSO₂alkenyl, —NRSO₂substituted alkenyl, —NRSO₂alkynyl,—NRSO₂substituted alkynyl, —NRSO₂aryl, —NRSO₂substituted aryl,—NRSO₂heteroaryl, —NRSO₂substituted heteroaryl, —NRSO₂heterocyclic, and—NRSO₂substituted heterocyclic where R is hydrogen or alkyl and whereinalkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl,substituted alkynyl, cycloalkyl, substituted cycloalkyl, aryl,substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic andsubstituted heterocyclic are as defined herein.

[0155] “Aminocarbonyloxy” refers to the groups —NRC(O)O-alkyl,—NRC(O)O-substituted alkyl, —NRC(O)O-alkenyl, —NRC(O)O-substitutedalkenyl, —NRC(O)O-alkynyl, —NRC(O)O-substituted alkynyl,—NRC(O)O-cycloalkyl, —NRC(O)O-substituted cycloalkyl, —NRC(O)O-aryl,—NRC(O)O-substituted aryl, —NRC(O)O-heteroaryl, —NRC(O)O-substitutedheteroaryl, —NRC(O)O-heterocyclic, and —NRC(O)O-substituted heterocyclicwhere R is hydrogen or alkyl and wherein alkyl, substituted alkyl,alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl,substituted cycloalkyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

[0156] “Aminosulfonyloxy” refers to the groups —NRSO₂O-alkyl,—NRSO₂O-substituted alkyl, —NRSO₂O-alkenyl, —NRSO₂O-substituted alkenyl,—NRSO₂O-alkynyl, —NRSO₂O-substituted alkynyl, —NRSO₂O-cycloalkyl,—NRSO₂O-substituted cycloalkyl, —NRSO₂O-aryl, —NRSO₂O-substituted aryl,—NRSO₂O-heteroaryl, —NRSO₂O-substituted heteroaryl,—NRSO₂O-heterocyclic, and —NRSO₂O-substituted heterocyclic where R ishydrogen or alkyl and wherein alkyl, substituted alkyl, alkenyl,substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl,substituted cycloalkyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

[0157] “Oxycarbonylamino” refers to the groups —OC(O)NH₂, —OC(O)NRR,—OC(O)NR-alkyl, —OC(O)NR-substituted alkyl, —OC(O)NR-alkenyl,—OC(O)NR-substituted alkenyl, —OC(O)NR-alkynyl, —OC(O)NR-substitutedalkynyl, —OC(O)NR-cycloalkyl, —OC(O)NR-substituted cycloalkyl,—OC(O)NR-aryl, —OC(O)NR-substituted aryl, —OC(O)NR-heteroaryl,—OC(O)NR-substituted heteroaryl, —OC(O)NR-heterocyclic, and—OC(O)NR-substituted heterocyclic where R is hydrogen, alkyl or whereeach R is joined to form, together with the nitrogen atom a heterocyclicor substituted heterocyclic ring and wherein alkyl, substituted alkyl,alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl,substituted cycloalkyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

[0158] “Oxythiocarbonylamino” refers to the groups —OC(S)NH₂, —OC(S)NRR,—OC(S)NR-alkyl, —OC(S)NR-substituted alkyl, —OC(S)NR-alkenyl,—OC(S)NR-substituted alkenyl, —OC(S)NR-alkynyl, —OC(S)NR-substitutedalkynyl, —OC(S)NR-cycloalkyl, —OC(S)NR-substituted cycloalkyl,—OC(S)NR-aryl, —OC(S)NR-substituted aryl, —OC(S)NR-heteroaryl,—OC(S)NR-substituted heteroaryl, —OC(S)NR-heterocyclic, and—OC(S)NR-substituted heterocyclic where R is hydrogen, alkyl or whereeach R is joined to form together with the nitrogen atom a heterocyclicor substituted heterocyclic ring and wherein alkyl, substituted alkyl,alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl,substituted cycloalkyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

[0159] “Oxysulfonylamino” refers to the groups —OSO₂NH₂, —OSO₂NRR,—OSO₂NR-alkyl, —OSO₂NR-substituted alkyl, —OSO₂NR-alkenyl,—OSO₂NR-substituted alkenyl, —OSO₂NR-alkynyl, —OSO₂NR-substitutedalkynyl, —OSO₂NR-cycloalkyl, —OSO₂NR-substituted cycloalkyl,—OSO₂NR-aryl, —OSO₂NR-substituted aryl, —OSO₂NR-heteroaryl,—OSO₂NR-substituted heteroaryl, —OSO₂NR-heterocyclic, and—OSO₂NR-substituted heterocyclic where R is hydrogen, alkyl or whereeach R is joined to form, together with the nitrogen atom a heterocyclicor substituted heterocyclic ring and wherein alkyl, substituted alkyl,alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl,substituted cycloalkyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

[0160] “Aminocarbonylamino” refers to the groups—NRC(O)NRR,—NRC(O)NR-alkyl, —NRC(O)NR-substituted alkyl,—NRC(O)NR-alkenyl, —NRC(O)NR-substituted alkenyl, —NRC(O)NR-alkynyl,—NRC(O)NR-substituted alkynyl, —NRC(O)NR-aryl, —NRC(O)NR-substitutedaryl, —NRC(O)NR-cycloalkyl, —NRC(O)NR-substituted cycloalkyl,—NRC(O)NR-heteroaryl, and —NRC(O)NR-substituted heteroaryl,—NRC(O)NR-heterocyclic, and —NRC(O)NR-substituted heterocyclic whereeach R is independently hydrogen, alkyl or where each R is joined toform together with the nitrogen atom a heterocyclic or substitutedheterocyclic ring as well as where one of the amino groups is blocked byconventional blocking groups such as Boc, Cbz, formyl, and the like andwherein alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl,substituted alkynyl, cycloalkyl, substituted cycloalkyl, aryl,substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic andsubstituted heterocyclic are as defined herein.

[0161] “Aminothiocarbonylamino” refers to the groups —NRC(S)NRR,—NRC(S)NR-alkyl, —NRC(S)NR-substituted alkyl, —NRC(S)NR-alkenyl,—NRC(S)NR-substituted alkenyl, —NRC(S)NR-alkynyl, —NRC(S)NR-substitutedalkynyl, —NRC(S)NR-aryl, —NRC(S)NR-substituted aryl,—NRC(S)NR-cycloalkyl, —NRC(S)NR-substituted cycloalkyl,—NRC(S)NR-heteroaryl, and —NRC(S)NR-substituted heteroaryl,—NRC(S)NR-heterocyclic, and —NRC(S)NR-substituted heterocyclic whereeach R is independently hydrogen, alkyl or where each R is joined toform together with the nitrogen atom a heterocyclic or substitutedheterocyclic ring as well as where one of the amino groups is blocked byconventional blocking groups such as Boc, Cbz, formyl, and the like andwherein alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl,substituted alkynyl, cycloalkyl, substituted cycloalkyl, aryl,substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic andsubstituted heterocyclic are as defined herein.

[0162] “Aminosulfonylamino” refers to the groups —NRSO₂NRR,—NRSO₂NR-alkyl, —NRSO₂NR-substituted alkyl, —NRSO₂NR-alkenyl,—NRSO₂NR-substituted alkenyl, —NRSO₂NR-alkynyl, —NRSO₂NR-substitutedalkynyl, —NRSO₂NR-aryl, —NRSO₂NR-substituted aryl, —NRSO₂NR-cycloalkyl,—NRSO₂NR-substituted cycloalkyl, —NRSO₂NR-heteroaryl, and—NRSO₂NR-substituted heteroaryl, —NRSO₂NR-heterocyclic, and—NRSO₂NR-substituted heterocyclic, where each R is independentlyhydrogen, alkyl or where each R is joined to form together with thenitrogen atom a heterocyclic or substituted heterocyclic ring as well aswhere one of the amino groups is blocked by conventional blocking groupssuch as Boc, Cbz, formyl, and the like and wherein alkyl, substitutedalkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl,cycloalkyl, substituted cycloalkyl, aryl, substituted aryl, heteroaryl,substituted heteroaryl, heterocyclic and substituted heterocyclic are asdefined herein.

[0163] “Aryl” or “Ar” refers to an unsaturated aromatic carbocyclicgroup of from 6 to 14 carbon atoms having a single ring (e.g., phenyl)or multiple condensed rings (e.g., naphthyl or anthryl) which condensedrings may or may not be aromatic (e.g., 2-benzoxazolinone,2H-1,4-benzoxazin-3(4H)-one-7yl, and the like). Preferred aryls includephenyl and naphthyl.

[0164] Substituted aryl refers to aryl groups which are substituted withfrom 1 to 3 substituents selected from the group consisting of hydroxy,acyl, acylamino, thiocarbonylamino, acyloxy, alkyl, substituted alkyl,alkoxy, substituted alkoxy, alkenyl, substituted alkenyl, alkynyl,substituted alkynyl, amidino, alkylamidino, thioamidino, amino,aminoacyl, aminocarbonyloxy, aminocarbonylamino, aminothiocarbonylamino,aryl, substituted aryl, aryloxy, substituted aryloxy, cycloalkoxy,substituted cycloalkoxy, heteroaryloxy, substituted heteroaryloxy,heterocyclyloxy, substituted heterocyclyloxy, carboxyl, carboxylalkyl,carboxyl-substituted alkyl, carboxyl-cycloalkyl, carboxyl-substitutedcycloalkyl, carboxylaryl, carboxyl-substituted aryl, carboxylheteroaryl,carboxyl-substituted heteroaryl, carboxylheterocyclic,carboxyl-substituted heterocyclic, carboxylamido, cyano, thiol,thioalkyl, substituted thioalkyl, thioaryl, substituted thioaryl,thioheteroaryl, substituted thioheteroaryl, thiocycloalkyl, substitutedthiocycloalkyl, thioheterocyclic, substituted thioheterocyclic,cycloalkyl, substituted cycloalkyl, guanidino, guanidinosulfone, halo,nitro, heteroaryl, substituted heteroaryl, heterocyclic, substitutedheterocyclic, cycloalkoxy, substituted cycloalkoxy, heteroaryloxy,substituted heteroaryloxy, heterocyclyloxy, substituted heterocyclyloxy,oxycarbonylamino, oxythiocarbonylamino, —S(O)₂-alkyl, —S(O)₂-substitutedalkyl, —S(O)₂-cycloalkyl, —S(O)₂-substituted cycloalkyl, —S(O)₂-alkenyl,—(O)₂-substituted alkenyl, —S(O)₂-aryl, —S(O)₂-substituted aryl,—S(O)₂-heteroaryl, —S(O)₂-substituted heteroaryl, —S(O)₂-heterocyclic,—S(O)₂-substituted heterocyclic, —OS(O)₂-alkyl, —OS(O)₂-substitutedalkyl, —OS(O)₂-aryl, —OS(O)₂-substituted aryl, —OS(O)₂-heteroaryl,—OS(O)₂-substituted heteroaryl, —OS(O)₂-heterocyclic,—OS(O)₂-substituted heterocyclic, —OSO₂—NRR where R is hydrogen oralkyl, —NRS(O)₂-alkyl, —NRS(O)₂-substituted alkyl, —NRS(O)₂-aryl,—NRS(O)₂-substituted aryl, —NRS(O)₂-heteroaryl, —NRS(O)₂-substitutedheteroaryl, —NRS(O)₂-heterocyclic, —NRS(O)₂-substituted heterocyclic,—NRS(O)₂—NR-alkyl, —NRS(O)₂—NR-substituted alkyl, —NRS(O)₂—NR-aryl,—NRS(O)₂—NR-substituted aryl, —NRS(O)₂—NR-heteroaryl,—NRS(O)₂—NR-substituted heteroaryl, —NRS(O)₂—NR-heterocyclic,—NRS(O)₂—NR-substituted heterocyclic where R is hydrogen or alkyl, mono-and di-alkylamino, mono- and di-(substituted alkyl)amino, mono- anddi-arylamino, mono- and di-substituted arylamino, mono- anddi-heteroarylamino, mono- and di-substituted heteroarylamino, mono- anddi-heterocyclic amino, mono- and di-substituted heterocyclic amino,unsymmetric di-substituted amines having different substituents selectedfrom alkyl, substituted alkyl, aryl, substituted aryl, heteroaryl,substituted heteroaryl, heterocyclic and substituted heterocyclic andamino groups on the substituted aryl blocked by conventional blockinggroups such as Boc, Cbz, formyl, and the like or substituted with—SO₂NRR where R is hydrogen or alkyl.

[0165] “Aryloxy” refers to the group aryl-O— which includes, by way ofexample, phenoxy, naphthoxy, and the like.

[0166] “Substituted aryloxy” refers to substituted aryl-O— groups.

[0167] “Aryloxyaryl” refers to the group-aryl-O-aryl.

[0168] “substituted aryloxyaryl” refers to aryloxyaryl groupssubstituted with from 1 to 3 substituents on either or both aryl ringsselected from the group consisting of hydroxy, acyl, acylamino,thiocarbonylamino, acyloxy, alkyl, substituted alkyl, alkoxy,substituted alkoxy, alkenyl, substituted alkenyl, alkynyl, substitutedalkynyl, amidino, alkylamidino, thioamidino, amino, aminoacyl,aminocarbonyloxy, aminocarbonylamino, aminothiocarbonylamino, aryl,substituted aryl, aryloxy, substituted aryloxy, cycloalkoxy, substitutedcycloalkoxy, heteroaryloxy, substituted heteroaryloxy, heterocyclyloxy,substituted heterocyclyloxy, carboxyl, carboxylalkyl,carboxyl-substituted alkyl, carboxyl-cycloalkyl, carboxyl-substitutedcycloalkyl, carboxylaryl, carboxyl-substituted aryl, carboxylheteroaryl,carboxyl-substituted heteroaryl, carboxylheterocyclic,carboxyl-substituted heterocyclic, carboxylamido, cyano, thiol,thioalkyl, substituted thioalkyl, thioaryl, substituted thioaryl,thioheteroaryl, substituted thioheteroaryl, thiocycloalkyl, substitutedthiocycloalkyl, thioheterocyclic, substituted thioheterocyclic,cycloalkyl, substituted cycloalkyl, guanidino, guanidinosulfone, halo,nitro, heteroaryl, substituted heteroaryl, heterocyclic, substitutedheterocyclic, cycloalkoxy, substituted cycloalkoxy, heteroaryloxy,substituted heteroaryloxy, heterocyclyloxy, substituted heterocyclyloxy,oxycarbonylamino, oxythiocarbonylamino, —S(O)₂-alkyl, —S(O)₂-substitutedalkyl, —S(O)₂-cycloalkyl, —S(O)₂-substituted cycloalkyl, —S(O)₂-alkenyl,—S(O)₂-substituted alkenyl, —S(O)₂-aryl, —S(O)₂-substituted aryl,—S(O)₂-heteroaryl, —S(O)₂-substituted heteroaryl, —S(O)₂-heterocyclic,—S(O)₂-substituted heterocyclic, —OS(O)₂-alkyl, —OS(O)₂-substitutedalkyl, —OS(O)₂-aryl, —OS(O)₂-substituted aryl, —OS(O)₂-heteroaryl,—OS(O)₂-substitutedheteroaryl, —OS(O)₂-heterocyclic, —OS(O)₂-substitutedheterocyclic, —OSO₂—NRR where R is hydrogen or alkyl, —NRS(O)₂-alkyl,—NRS(O)₂-substituted alkyl, —NRS(O)₂-aryl, —NRS(O)₂-substituted aryl,—NRS(O)₂-heteroaryl, —NRS(O)₂-substituted heteroaryl,—NRS(O)₂-heterocyclic, —NRS(O)₂-substituted heterocyclic,—NRS(O)₂—NR-alkyl, —NRS(O)₂—NR-substituted alkyl, —NRS(O)₂—NR-aryl,—NRS(O)₂—NR-substituted aryl, —NRS(O)₂—NR-heteroaryl,—NRS(O)₂—NR-substituted heteroaryl, —NRS(O)₂—NR-heterocyclic,—NRS(O)₂—NR-substituted heterocyclic where R is hydrogen or alkyl, mono-and di-alkylamino, mono- and di-(substituted alkyl)amino, mono- anddi-arylamino, mono- and di-substituted arylamino, mono- anddi-heteroarylamino, mono- and di-substituted heteroarylamino, mono- anddi-heterocyclic amino, mono- and di-substituted heterocyclic amino,unsymmetric di-substituted amines having different substituents selectedfrom alkyl, substituted alkyl, aryl, substituted aryl, heteroaryl,substituted heteroaryl, heterocyclic and substituted heterocyclic andamino groups on the substituted aryl blocked by conventional blockinggroups such as Boc, Cbz, formyl, and the like or substituted with—SO₂NRR where R is hydrogen or alkyl.

[0169] “Cycloalkyl” refers to cyclic alkyl groups of from 3 to 8 carbonatoms having a single cyclic ring including, by way of example,cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclooctyl and thelike. Excluded from this definition are multi-ring alkyl groups such asadamantanyl, etc.

[0170] “Cycloalkenyl” refers to cyclic alkenyl groups of from 3 to 8carbon atoms having single or multiple unsaturation but which are notaromatic.

[0171] “Substituted cycloalkyl” and “substituted cycloalkenyl” refer toa cycloalkyl and cycloalkenyl groups, preferably of from 3 to 8 carbonatoms, having from 1 to 5 substituents selected from the groupconsisting of oxo(=O), thioxo(=S), alkoxy, substituted alkoxy, acyl,acylamino, thiocarbonylamino, acyloxy, amino, amidino, alkylamidino,thioamidino, aminoacyl, aminocarbonylamino, aminothiocarbonylamino,aminocarbonyloxy, aryl, substituted aryl, aryloxy, substituted aryloxy,aryloxyaryl, substituted aryloxyaryl, halogen, hydroxyl, cyano, nitro,carboxyl, carboxylalkyl, carboxyl-substituted alkyl,carboxyl-cycloalkyl, carboxyl-substituted cycloalkyl, carboxylaryl,carboxyl-substituted aryl, carboxylheteroaryl, carboxyl-substitutedheteroaryl, carboxylheterocyclic, carboxyl-substituted heterocyclic,cycloalkyl, substituted cycloalkyl, guanidino, guanidinosulfone, thiol,thioalkyl, substituted thioalkyl, thioaryl, substituted thioaryl,thiocycloalkyl, substituted thiocycloalkyl, thioheteroaryl, substitutedthioheteroaryl, thioheterocyclic, substituted thioheterocyclic,heteroaryl, substituted heteroaryl, heterocyclic, substitutedheterocyclic, cycloalkoxy, substituted cycloalkoxy, heteroaryloxy,substituted heteroaryloxy, heterocyclyloxy, substituted heterocyclyloxy,oxycarbonylamino, oxythiocarbonylamino, —OS(O)₂-alkyl,—OS(O)₂-substituted alkyl, —OS(O)₂-aryl, —OS(O)₂-substituted aryl,—OS(O)₂-heteroaryl, —OS(O)₂-substituted heteroaryl,—OS(O)₂-heterocyclic, —OS(O)₂-substituted heterocyclic, —OSO₂—NRR whereR is hydrogen or alkyl, —NRS(O)₂-alkyl, —NRS(O)₂-substituted alkyl,—NRS(O)₂-aryl, —NRS(O)₂-substituted aryl, —NRS(O)₂-heteroaryl,—NRS(O)₂-substituted heteroaryl, —NRS(O)₂-heterocyclic,—NRS(O)₂-substituted heterocyclic, —NRS(O)₂—NR-alkyl,—NRS(O)₂—NR-substituted alkyl, —NRS(O)₂—NR-aryl, —NRS(O)₂—NR-substitutedaryl, —NRS(O)₂—NR-heteroaryl, —NRS(O)₂—NR-substituted heteroaryl,—NRS(O)₂—NR-heterocyclic, —NRS(O)₂—NR-substituted heterocyclic where Ris hydrogen or alkyl, mono- and di-alkylamino, mono- and di-(substitutedalkyl)amino, mono- and di-arylamino, mono- and di-substituted arylamino,mono- and di-heteroarylamino, mono- and di-substituted heteroarylamino,mono- and di-heterocyclic amino, mono- and di-substituted heterocyclicamino, unsymmetric di-substituted amines having different substituentsselected from alkyl, substituted alkyl, aryl, substituted aryl,heteroaryl, substituted heteroaryl, heterocyclic and substitutedheterocyclic and substituted alkynyl groups having amino groups blockedby conventional blocking groups such as Boc, Cbz, formyl, and the likeor alkynyl/substituted alkynyl groups substituted with —SO₂-alkyl,—SO₂-substituted alkyl, —SO₂-alkenyl, —SO₂-substituted alkenyl,—SO₂-cycloalkyl, —SO₂-substituted cycloalkyl, —SO₂-aryl,—SO₂-substituted aryl, —SO₂-heteroaryl, —SO₂-substituted heteroaryl,—SO₂-heterocyclic, —SO₂-substituted heterocyclic and —SO₂NRR where R ishydrogen or alkyl.

[0172] “Cycloalkoxy” refers to —O-cycloalkyl groups.

[0173] “Substituted cycloalkoxy” refers to —O-substituted cycloalkylgroups.

[0174] “Cycloalkenoxy” refers to —O-cycloalkenyl groups.

[0175] “Substituted cycloalkenoxy” refers to —O-substituted cycloalkenylgroups.

[0176] “Guanidino” refers to the groups —NRC(═NR)NRR, —NRC(═NR)NR-alkyl,—NRC(═NR)NR-substituted alkyl, —NRC( ═NR)NR-alkenyl,—NRC(═NR)NR-substituted alkenyl, —NRC(═NR)NR-alkynyl,—NRC(═NR)NR-substituted alkynyl, —NRC(═NR)NR-aryl,—NRC(═NR)NR-substituted aryl, —NRC(═NR)NR-cycloalkyl,—NRC(═NR)NR-heteroaryl, —NRC(═NR)NR-substituted heteroaryl,—NRC(═NR)NR-heterocyclic, and —NRC(═NR)NR-substituted heterocyclic whereeach R is independently hydrogen and alkyl as well as where one of theamino groups is blocked by conventional blocking groups such as Boc,Cbz, formyl, and the like and wherein alkyl, substituted alkyl, alkenyl,substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl,substituted cycloalkyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

[0177] “Guanidinosulfone” refers to the groups —NRC(═NR)NRSO₂-alkyl,—NRC(═NR)NRSO₂-substituted alkyl, —NRC(═NR)NRSO₂-alkenyl,—NRC(═NR)NRSO₂-substituted alkenyl, —NRC(═NR)NRSO₂-alkynyl,—NRC(═NR)NRSO₂-substituted alkynyl, —NRC(═NR)NRSO₂-aryl,—NRC(═NR)NRSO₂-substituted aryl, —NRC(═NR)NRSO₂-cycloalkyl,—NRC(═NR)NRSO₂-substituted cycloalkyl, —NRC(═NR)NRSO₂-heteroaryl, and—NRC(═NR)NRSO₂-substituted heteroaryl, —NRC(═NR)NRSO₂-heterocyclic, and—NRC(═NR)NRSO₂-substituted heterocyclic where each R is independentlyhydrogen and alkyl and wherein alkyl, substituted alkyl, alkenyl,substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl,substituted cycloalkyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

[0178] “Halo” or “halogen” refers to fluoro, chloro, bromo and iodo andpreferably is either chloro or bromo.

[0179] “Heteroaryl” refers to an aromatic carbocyclic group of from 2 to10 carbon atoms and 1 to 4 heteroatoms selected from oxygen, nitrogenand sulfur within the ring or oxides thereof. Such heteroaryl groups canhave a single ring (e.g., pyridyl or furyl) or multiple condensed rings(e.g., indolizinyl or benzothienyl). Additionally, the heteroatoms ofthe heteroaryl group may be oxidized, i.e., to form pyridine N-oxides or1,1-dioxo-1,2,5-thiadiazoles and the like. Preferred heteroaryls includepyridyl, pyrrolyl, indolyl, furyl, pyridazinyl, pyrimidinyl, pyrazinyl,1-oxo-1,2,5-thiadiazolyl and 1,1-dioxo-1,2,5-thiadiazolyl. The term“heteroaryl having two nitrogen atoms in the heteroaryl ring” refers toa heteroaryl group having two, and only two, nitrogen atoms in theheteroaryl ring and optionally containing 1 or 2 other heteroatoms inthe heteroaryl ring, such as oxygen or sulfur

[0180] “Substituted heteroaryl” refers to heteroaryl groups which aresubstituted with from 1 to 3 substituents selected from the groupconsisting of hydroxy, acyl, acylamino, thiocarbonylamino, acyloxy,alkyl, substituted alkyl, alkoxy, substituted alkoxy, alkenyl,substituted alkenyl, alkynyl, substituted alkynyl, amidino,alkylamidino, thioamidino, amino, aminoacyl, aminocarbonyloxy,aminocarbonylamino, aminothiocarbonylamino, aryl, substituted aryl,aryloxy, substituted aryloxy, cycloalkoxy, substituted cycloalkoxy,heteroaryloxy, substituted heteroaryloxy, heterocyclyloxy, substitutedheterocyclyloxy, carboxyl, carboxylalkyl, carboxyl-substituted alkyl,carboxyl-cycloalkyl, carboxyl-substituted cycloalkyl, carboxylaryl,carboxyl-substituted aryl, carboxylheteroaryl, carboxyl-substitutedheteroaryl, carboxylheterocyclic, carboxyl-substituted heterocyclic,carboxylamido, cyano, thiol, thioalkyl, substituted thioalkyl, thioaryl,substituted thioaryl, thioheteroaryl, substituted thioheteroaryl,thiocycloalkyl, substituted thiocycloalkyl, thioheterocyclic,substituted thioheterocyclic, cycloalkyl, substituted cycloalkyl,guanidino, guanidinosulfone, halo, nitro, heteroaryl, substitutedheteroaryl, heterocyclic, substituted heterocyclic, cycloalkoxy,substituted cycloalkoxy, heteroaryloxy, substituted heteroaryloxy,heterocyclyloxy, substituted heterocyclyloxy, oxycarbonylamino,oxythiocarbonylamino, —S(O)₂-alkyl, —S(O)₂-substituted alkyl,—S(O)₂-cycloalkyl, —S(O)₂-substituted cycloalkyl, —S(O)₂-alkenyl,—S(O)₂-substituted alkenyl, —S(O)₂-aryl, —S(O)₂-substituted aryl,—S(O)₂-heteroaryl, —S(O)₂-substituted heteroaryl, —S(O)₂-heterocyclic,—S(O)₂-substituted heterocyclic, —OS(O)₂-alkyl, —OS(O)₂-substitutedalkyl, —OS(O)₂-aryl, —OS(O)₂-substituted aryl, —OS(O)₂-heteroaryl,—OS(O)₂-substituted heteroaryl, —OS(O)₂-heterocyclic,—OS(O)₂-substituted heterocyclic, —OSO₂—NRR where R is hydrogen oralkyl, —NRS(O)₂-alkyl, —NRS(O)₂-substituted alkyl, —NRS(O)₂-aryl,—NRS(O)₂-substituted aryl, —NRS(O)₂-heteroaryl, —NRS(O)₂-substitutedheteroaryl, —NRS(O)₂-heterocyclic, —NRS(O)₂-substituted heterocyclic,—NRS(O)₂—NR-alkyl, —NRS(O)₂—NR-substituted alkyl, —NRS(O)₂—NR-aryl,—NRS(O)₂—NR-substituted aryl, —NRS(O)₂—NR-heteroaryl,—NRS(O)₂—NR-substituted heteroaryl, —NRS(O)₂—NR-heterocyclic,—NRS(O)₂—NR-substituted heterocyclic where R is hydrogen or alkyl, mono-and di-alkylamino, mono- and di-(substituted alkyl)amino, mono- anddi-arylamino, mono- and di-substituted arylamino, mono- anddi-heteroarylamino, mono- and di-substituted heteroarylamino, mono- anddi-heterocyclic amino, mono- and di-substituted heterocyclic amino,unsymmetric di-substituted amines having different substituents selectedfrom alkyl, substituted alkyl, aryl, substituted aryl, heteroaryl,substituted heteroaryl, heterocyclic and substituted heterocyclic andamino groups on the substituted aryl blocked by conventional blockinggroups such as Boc, Cbz, formyl, and the like or substituted with—SO₂NRR where R is hydrogen or alkyl.

[0181] “Nitrogen containing heteroaryl” refers to a heteroary ring asdefined above that contains at least one nitrogen atom in the ring. Suchheteroaryl groups can have a single ring (e.g., pyridyl) or multiplecondensed rings (e.g., indolizinyl). Additionally, the heteroatoms ofthe heteroaryl group may be oxidized, i.e., to form pyridine N-oxides or1,1-dioxo-1,2,5-thiadiazoles and the like. Preferred nitrogen containingheteroaryls include pyridyl, pyrrolyl, indolyl, pyridazinyl,pyrimidinyl, pyrazinyl, 1-oxo-1,2,5-thiadiazolyl and1,1-dioxo-1,2,5-thiadiazolyl. The term “heteroaryl having two nitrogenatoms in the heteroaryl ring” refers to a heteroaryl group having two,and only two, nitrogen atoms in the heteroaryl ring and optionallycontaining 1 or 2 other heteroatoms in the heteroaryl ring, such asoxygen or sulfur

[0182] “Nitrogen containing substituted heteroaryl” refers to heteroarylgroups as defined above which are substituted with from 1 to 3substituents selected from the group consisting of hydroxy, acyl,acylamino, thiocarbonylamino, acyloxy, alkyl, substituted alkyl, alkoxy,substituted alkoxy, alkenyl, substituted alkenyl, alkynyl, substitutedalkynyl, amidino, alkylamidino, thioamidino, amino, aminoacyl,aminocarbonyloxy, aminocarbonylamino, aminothiocarbonylamino, aryl,substituted aryl, aryloxy, substituted aryloxy, cycloalkoxy, substitutedcycloalkoxy, heteroaryloxy, substituted heteroaryloxy, heterocyclyloxy,substituted heterocyclyloxy, carboxyl, carboxylalkyl,carboxyl-substituted alkyl, carboxyl-cycloalkyl, carboxyl-substitutedcycloalkyl, carboxylaryl, carboxyl-substituted aryl, carboxylheteroaryl,carboxyl-substituted heteroaryl, carboxylheterocyclic,carboxyl-substituted heterocyclic, carboxylamido, cyano, thiol,thioalkyl, substituted thioalkyl, thioaryl, substituted thioaryl,thioheteroaryl, substituted thioheteroaryl, thiocycloalkyl, substitutedthiocycloalkyl, thioheterocyclic, substituted thioheterocyclic,cycloalkyl, substituted cycloalkyl, guanidino, guanidinosulfone, halo,nitro, heteroaryl, substituted heteroaryl, heterocyclic, substitutedheterocyclic, cycloalkoxy, substituted cycloalkoxy, heteroaryloxy,substituted heteroaryloxy, heterocyclyloxy, substituted heterocyclyloxy,oxycarbonylamino, oxythiocarbonylamino, —S(O)₂-alkyl, —S(O)₂-substitutedalkyl, —S(O)₂-cycloalkyl, —S(O)₂-substituted cycloalkyl, —S(O)₂-alkenyl,—S(O)₂-substituted alkenyl, —S(O)₂-aryl, —S(O)₂-substituted aryl,—S(O)₂-heteroaryl, —S(O)₂-substituted heteroaryl, —S(O)₂-heterocyclic,—S(O)₂-substituted heterocyclic, —OS(O)₂-alkyl, —OS(O)₂-substitutedalkyl, —S(O)₂-aryl, —OS(O)₂-substituted aryl, —OS(O)₂-heteroaryl,—OS(O)₂-substituted heteroaryl, —OS(O)₂-heterocyclic,—OS(O)₂-substituted heterocyclic, —OSO₂—NRR where R is hydrogen oralkyl, —NRS(O)₂-alkyl, —NRS(O)₂-substituted alkyl, —NRS(O)₂-aryl,—NRS(O)₂-substituted aryl, —NRS(O)₂-heteroaryl, —NRS(O)₂-substitutedheteroaryl, —NRS(O)₂-heterocyclic, —NRS(O)₂-substituted heterocyclic,—NRS(O)₂—NR-alkyl, —NRS(O)₂—NR-substituted alkyl, —NRS(O)₂—NR-aryl,—NRS(O)₂—NR-substituted aryl, —NRS(O)₂—NR-heteroaryl,—NRS(O)₂—NR-substituted heteroaryl, —NRS(O)₂—NR-heterocyclic,—NRS(O)₂—NR-substituted heterocyclic where R is hydrogen or alkyl, mono-and di-alkylamino, mono- and di-(substituted alkyl)amino, mono- anddi-arylamino, mono- and di-substituted arylamino, mono- anddi-heteroarylamino, mono- and di-substituted heteroarylamino, mono- anddi-heterocyclic amino, mono- and di-substituted heterocyclic amino,unsymmetric di-substituted amines having different substituents selectedfrom alkyl, substituted alkyl, aryl, substituted aryl, heteroaryl,substituted heteroaryl, heterocyclic and substituted heterocyclic andamino groups on the substituted aryl blocked by conventional blockinggroups such as Boc, Cbz, formyl, and the like or substituted with—SO₂NRR where R is hydrogen or alkyl.

[0183] “Heteroaryloxy” refers to the group —O-heteroaryl and“substituted heteroaryloxy” refers to the group —O-substitutedheteroaryl.

[0184] “Heterocycle” or “heterocyclic” refers to a saturated orunsaturated group having a single ring or multiple condensed rings, from1 to 10 carbon atoms and from 1 to 4 hetero atoms selected fromnitrogen, sulfur or oxygen within the ring wherein, in fused ringsystems, one or more of the rings can be aryl or heteroaryl.

[0185] “Substituted heterocyclic” refers to heterocycle groups which aresubstituted with from 1 to 3 substituents selected from the groupconsisting of oxo(=O), thioxo(=S), alkoxy, substituted alkoxy, acyl,acylamino, thiocarbonylamino, acyloxy, amino, amidino, alkylamidino,thioamidino, aminoacyl, aminocarbonylamino, aminothiocarbonylamino,aminocarbonyloxy, aryl, substituted aryl, aryloxy, substituted aryloxy,aryloxyaryl, substituted aryloxyaryl, halogen, hydroxyl, cyano, nitro,carboxyl, carboxylalkyl, carboxyl-substituted alkyl,carboxyl-cycloalkyl, carboxyl-substituted cycloalkyl, carboxylaryl,carboxyl-substituted aryl, carboxylheteroaryl, carboxyl-substitutedheteroaryl, carboxylheterocyclic, carboxyl-substituted heterocyclic,cycloalkyl, substituted cycloalkyl, guanidino, guanidinosulfone, thiol,thioalkyl, substituted thioalkyl, thioaryl, substituted thioaryl,thiocycloalkyl, substituted thiocycloalkyl, thioheteroaryl, substitutedthioheteroaryl, thioheterocyclic, substituted thioheterocyclic,heteroaryl, substituted heteroaryl, heterocyclic, substitutedheterocyclic, cycloalkoxy, substituted cycloalkoxy, heteroaryloxy,substituted heteroaryloxy, heterocyclyloxy, substituted heterocyclyloxy,oxycarbonylamino, oxythiocarbonylamino,—OS(O)₂-alkyl,—OS(O)₂-substituted alkyl, —OS(O)₂-aryl,—OS(O)₂-substituted aryl, —OS(O)₂-heteroaryl, —OS(O)₂-substitutedheteroaryl, —OS(O)₂-heterocyclic, —OS(O)₂-substituted heterocyclic,—OSO₂—NRR where R is hydrogen or alkyl, —NRS(O)₂-alkyl,—NRS(O)₂-substituted alkyl, —NRS(O)₂-aryl, —NRS(O)₂-substituted aryl,—NRS(O)₂-heteroaryl, —NRS(O)₂-substituted heteroaryl,—NRS(O)₂-heterocyclic, —NRS(O)₂-substituted heterocyclic,—NRS(O)₂—NR-alkyl, —NRS(O)₂—NR-substituted alkyl, —NRS(O)₂—NR-aryl,—NRS(O)₂—NR-substituted aryl, —NRS(O)₂—NR-heteroaryl,—NRS(O)₂—NR-substituted heteroaryl, —NRS(O)₂—NR-heterocyclic,—NRS(O)₂—NR-substituted heterocyclic where R is hydrogen or alkyl, mono-and di-alkylamino, mono- and di-(substituted alkyl)amino, mono- anddi-arylamino, mono- and di-substituted arylamino, mono- anddi-heteroarylamino, mono- and di-substituted heteroarylamino, mono- anddi-heterocyclic amino, mono- and di-substituted heterocyclic amino,unsymmetric di-substituted amines having different substituents selectedfrom alkyl, substituted alkyl, aryl, substituted aryl, heteroaryl,substituted heteroaryl, heterocyclic and substituted heterocyclic andsubstituted alkynyl groups having amino groups blocked by conventionalblocking groups such as Boc, Cbz, formyl, and the like oralkynyl/substituted alkynyl groups substituted with —SO₂-alkyl,—SO₂-substituted alkyl, —SO₂-alkenyl, —SO₂-substituted alkenyl,—SO₂-cycloalkyl, —SO₂-substituted cycloalkyl, —SO₂-aryl,—SO₂-substituted aryl, —SO₂-heteroaryl, —SO₂-substituted heteroaryl,—SO₂-heterocyclic, —SO₂-substituted heterocyclic and —SO₂NRR where R ishydrogen or alkyl.

[0186] Examples of heterocycles and heteroaryls include, but are notlimited to, azetidine, pyrrole, imidazole, pyrazole, pyridine, pyrazine,pyrimidine, pyridazine, indolizine, isoindole, indole, dihydroindole,indazole, purine, quinolizine, isoquinoline, quinoline, phthalazine,naphthylpyridine, quinoxaline, quinazoline, cinnoline, pteridine,carbazole, carboline, phenanthridine, acridine, phenanthroline,isothiazole, phenazine, isoxazole, phenoxazine, phenothiazine,imidazolidine, imidazoline, piperidine, piperazine, indoline,phthalimide, 1,2,3,4-tetrahydroisoquinoline,4,5,6,7-tetrahydrobenzo[b]thiophene, thiazole, thiazolidine, thiophene,benzo[b]thiophene, morpholino, thiomorpholino, piperidinyl, pyrrolidine,tetrahydrofuranyl, and the like.

[0187] “Heterocyclyloxy” refers to the group —O-heterocyclic and“substituted heterocyclyloxy” refers to the group —O-substitutedheterocyclic.

[0188] “Thiol” refers to the group —SH.

[0189] “Thioalkyl” refers to the groups —S-alkyl

[0190] “Substituted thioalkyl” refers to the group —S-substituted alkyl.

[0191] “Thiocycloalkyl” refers to the groups —S-cycloalkyl.

[0192] “Substituted thiocycloalkyl” refers to the group —S-substitutedcycloalkyl.

[0193] “Thioaryl” refers to the group —S-aryl and “substituted thioaryl”refers to the group —S-substituted aryl.

[0194] “Thioheteroaryl” refers to the group —S-heteroaryl and“substituted thioheteroaryl” refers to the group —S-substitutedheteroaryl.

[0195] “Thioheterocyclic” refers to the group —S-heterocyclic and“substituted thioheterocyclic” refers to the group —S-substitutedheterocyclic.

[0196] “Pharmaceutically acceptable salt” refers to pharmaceuticallyacceptable salts of a compound of Formula (I) which salts are derivedfrom a variety of organic and inorganic counter ions well known in theart and include, by way of example only, sodium, potassium, calcium,magnesium, ammonium, tetraalkylammonium, and the like; and when themolecule contains a basic functionality, salts of organic or inorganicacids, such as hydrochloride, hydrobromide, tartrate, mesylate, acetate,maleate, oxalate and the like.

General Synthetic Scheme

[0197] The compounds of this invention can be prepared from readilyavailable starting materials using the following general methods andprocedures. It will be appreciated that where typical or preferredprocess conditions (i.e., reaction temperatures, times, mole ratios ofreactants, solvents, pressures, etc.) are given, other processconditions can also be used unless otherwise stated. Optimum reactionconditions may vary with the particular reactants or solvent used, butsuch conditions can be determined by one skilled in the art by routineoptimization procedures.

[0198] Additionally, as will be apparent to those skilled in the art,conventional protecting groups may be necessary to prevent certainfunctional groups from undergoing undesired reactions. Suitableprotecting groups for various functional groups as well as suitableconditions for protecting and deprotecting particular functional groupsare well known in the art. For example, numerous protecting groups aredescribed in T. W. Greene and G. M. Wuts, Protecting Groups in OrganicSynthesis, Second Edition, Wiley, N.Y., 1991, and references citedtherein.

[0199] Furthermore, the compounds of this invention will typicallycontain one or more chiral centers. Accordingly, if desired, suchcompounds can be prepared or isolated as pure stereoisomers, i.e., asindividual enantiomers or diastereomers, or as stereoisomer-enrichedmixtures. All such stereoisomers (and enriched mixtures) are includedwithin the scope of this invention, unless otherwise indicated. Purestereoisomers (or enriched mixtures) may be prepared using, for example,optically active starting materials or stereoselective reagentswell-known in the art. Alternatively, racemic mixtures of such compoundscan be separated using, for example, chiral column chromatography,chiral resolving agents and the like.

[0200] In a preferred method of synthesis, the compounds of thisinvention are prepared by coupling an amino acid derivative of theformula (a):

[0201] where HetAr is as defined herein and P¹ is a carboxylic acidprotecting group (such as an alkyl group, i.e. methyl, ethyl and thelike), with a suitably functionalized cyclic group (e.g., heteroaryl orheterocyclic intermediate). For example, such coupling reactions may beperformed by displacing a leaving group, such as chloro, bromo, iodo,tosyl and the like, from the cyclic intermediate, e.g., heteroaryl orheterocyclic intermediate, with the amino group of the amino acidderivative; or by reductive alkylation of the amino group of amino acidderivative with a carbonyl-functionalized intermediate. Such couplingreactions are well-known to those skilled in the art.

[0202] By way of illustration, the synthesis of a representativecompound of Formula (I) is shown in Scheme 1.

[0203] As shown in Scheme 1,5-nitrouracil, 1, (commercially availablefrom Aldrich Chemical Company, Milwaukee, Wis. USA) is treated withphosphorus oxychloride and N,N-dimethylaniline according to theprocedure described in Whittaker, J. Chem. Soc. 1951, 1565 to give1,3-dichloro-4-nitropyrimidine, 2.

[0204] 1,3-Dichloro-4-nitropyrimidine, 2, is then reacted with about onemolar equivalent of an amino acid derivative of the formula:H₂N—CH(CH₂HetAr)C(O)X where HetAr and X are as defined herein or X is—OP¹ where P¹ is a carboxylic acid protecting group, in the presence ofa trialkylamine, such as diisopropylethylamine (DIEA). Typically, thisreaction is conducted in an inert diluent, such as dichloromethane, at atemperature ranging from about 0° C. to about 10° C. for about 5 min. toabout 6 hours to afford intermediate 3.

[0205] The nitro group of intermediate 3 is then reduced using aconventional reducing agent, such as hydrogen and a palladium on carboncatalyst. When hydrogen and palladium on carbon are employed as thereducing agent, the chloro group of intermediate 3 is also removed. Thisreaction is typically conducted by contacting 3 with a Degussa-typepalladium on carbon catalyst (typically 20%) and excess sodiumbicarbonate in an inert diluent, such as methanol, under hydrogen(typically about 55 psi) for about 12 to 36 hours at ambient temperatureto afford amino intermediate 4.

[0206] Amino intermediate 4 is then reacted with a sulfonyl chloride ofthe formula: R⁵—S(O)₂—Cl, where R⁵ is as defined herein, to providesulfonamide intermediate 5. This reaction is typically conducted byreacting the amino intermediate 4 with at least one equivalent,preferably about 1.1 to about 2 equivalents, of the sulfonyl chloride inan inert diluent such as dichloromethane and the like. Generally, thereaction is conducted at a temperature ranging from about −70° C. toabout 40° C. for about 1 to about 24 hours. Preferably, this reaction isconducted in the presence of a suitable base to scavenge the acidgenerated during the reaction. Suitable bases include, by way ofexample, tertiary amines, such as triethylamine, diisopropylethylamine,N-methylmorpholine and the like. Alternatively, the reaction can beconducted under Schotten-Baumann-type conditions using aqueous alkali,such as sodium hydroxide and the like, as the base. Upon completion ofthe reaction, the resulting sulfonamide 5 is recovered by conventionalmethods including neutralization, extraction, precipitation,chromatography, filtration, and the like.

[0207] Other heteroaryl intermediates may also be employed in the abovedescribed reactions including, but not limited to,2-chloro-3-nitropyrazine (J. Med. Chem. 1984, 27, 1634);4-chloro-5-nitroimidazole (J. Chem. Soc. 1930, 268); and the like.

[0208] The amino acid derivatives (a) employed in the above reactionsare either known compounds or compounds that can be prepared from knowncompounds by conventional synthetic procedures. For example, amino acidderivatives can be prepared by reacting an organolithium reagent of theformula HetArCH₂Li wherein HetAr group is as defined in the Summary ofthe Invention with commercially available diethyloxalate (Aldrich,Milwaukee, Wis., USA). This reaction is typically conducted by treatingthe diethyloxalate with at least one equivalent of HetArCH₂Li in drypolar organic solvent such as tetrahydrofuran at <−70° C., preferably,−78° C. After addition is completed, the reaction mixture is thenallowed to warm to room temperature and stirred for about 18 to about 20hours. The resulting 3-(HetAr)-oxalate ester is then converted to thecorresponding oxime by following the procedure described in Negi et al,Synthesis, 1996, 8, 991. The oxime is then converted to3-(HetAr)-alanine ethyl ester by the procedure described in Keck et al.Syn. Commun., 1979, 281-286.

[0209] Alternatively, a 3-(HetAr)alanine methyl ester can be preparedfrom N-benzyloxycarbonyldehydroalanine methyl ester by treating it withHetAr halide wherein HetAr is as defined in the Summary of theInvention, to provide 3-(HetAr)alaninedehydroalanine methyl ester asdescribed in Carlstrom and Frejd, J. Org. Chem., 1991, 56, 1289-1293.The 3-(HetAr)alanine-dehydroalanine methyl ester is then converted tothe desired 3-(HetAr)-alanine methyl ester by the procedure described inKeck et al. Syn. Commun., 1979, 281-286.

[0210] Examples of amino acid derivatives suitable for use in the abovereactions include, but are not limited to, L-N(-methyl)histidine methylester, L-(N-benzyl)histidine methyl ester, L-tryptophan methyl ester,β-(2-pyridyl)-L-alanine methyl ester, β-(3-pyridyl)-L-alanine methylester, β-(4-pyridyl)-L-alanine methyl ester, β-(2-thiazolyl)-D,L-alaninemethyl ester, β-(1,2,4-triazol-3-yl)-D,L-alanine methyl ester,β-[2-(2,6-dimethoxyphenyl)pyridyl)-D,L-alanine methyl ester,β-[2-(2,6-dimethoxyphenyl)pyridazinyl)-D,L-alanine methyl ester, 62-[2-(2,6-dimethoxyphenyl)pyridinyl)-D,L-alanine methyl ester,β-[2-(2,6-dimethoxyphenyl)pyrazinyl)-D,L-alanine methyl ester, and thelike. If desired, of course, other esters or amides of theabove-described compounds may also be employed. Synthesis of compoundssuch as β-[2-(2,6-dimethoxyphenyl)pyridyl)-D,L-alanine methyl ester,β-[2-(2,6-dimethoxyphenyl)pyridazinyl)-D,L-alanine methyl ester,β-[2-(2,6-dimethoxyphenyl)pyrimidinyl)-D,L-alanine methyl ester, andβ-[2-(2,6-dimethoxyphenyl)pyrazinyl)-D,L-alanine methyl ester isdescribed in more detail below.

[0211] The sulfonyl chlorides employed in the above reaction are alsoeither known compounds or compounds that can be prepared from knowncompounds by conventional synthetic procedures. Such compounds aretypically prepared from the corresponding sulfonic acid, i.e., fromcompounds of the formula R⁵—SO₃H where R⁵ is as defined above, usingphosphorous trichloride and phosphorous pentachloride. This reaction isgenerally conducted by contacting the sulfonic acid with about 2 to 5molar equivalents of phosphorous trichloride and phosphorouspentachloride, either neat or in an inert solvent, such asdichloromethane, at temperature in the range of about 0° C. to about 80°C. for about 1 to about 48 hours to afford the sulfonyl chloride.Alternatively, the sulfonyl chloride can be prepared from thecorresponding thiol compound, i.e., from compounds of the formula R⁵—SHwhere R⁵ is as defined herein, by treating the thiol with chlorine (Cl₂)and water under conventional reaction conditions.

[0212] Examples of sulfonyl chlorides suitable for use in this inventioninclude, but are not limited to, methanesulfonyl chloride,2-propanesulfonyl chloride, 1-butanesulfonyl chloride, benzenesulfonylchloride, 1-naphthalene-sulfonyl chloride, 2-naphthalenesulfonylchloride, p-toluenesulfonyl chloride, α-toluenesulfonyl chloride,4-acetamidobenzene-sulfonyl chloride, 4-amidinobenzenesulfonyl chloride,4-tert-butyl-benzenesulfonyl chloride, 4-bromobenzenesulfonyl chloride,2-carboxybenzenesulfonyl chloride, 4-cyanobenzenesulfonyl chloride,3,4-dichlorobenzenesulfonyl chloride, 3,5-dichlorobenzenesulfonylchloride, 3,4-dimethoxybenzenesulfonyl chloride,3,5-ditrifluoromethylbenzenesulfonyl chloride, 4-fluorobenzenesulfonylchloride, 4-methoxybenzenesulfonyl chloride,2-methoxycarbonyl-benzenesulfonyl chloride, 4-methylamidobenzenesulfonylchloride, 4-nitrobenzenesulfonyl chloride, 4-thioamidobenzenesulfonylchloride, 4-trifluoromethylbenzenesulfonyl chloride,4-trifluoromethoxybenzenesulfonyl chloride,2,4,6-trimethylbenzenesulfonyl chloride, 2-phenylethanesulfonylchloride, 2-thiophenesulfonyl chloride, 5-chloro-2-thiophenesulfonylchloride, 2,5-dichloro-4-thiophenesulfonyl chloride, 2-thiazolesulfonylchloride, -methyl-4-thiazolesulfonyl chloride,1-methyl-4-imidazolesulfonyl chloride, 1-methyl-4-pyrazolesulfonylchloride, 5-chloro-1,3-dimethyl-4-pyrazole-sulfonyl chloride,3-pyridinesulfonyl chloride, 2-pyrimidinesulfonyl chloride and the like.If desired, a sulfonyl fluoride, sulfonyl bromide or sulfonic acidanhydride may be used in place of the sulfonyl chloride in the abovereaction to form the sulfonamide intermediate 5.

[0213] If desired, sulfonamide intermediate 5 can be alkylated at thesulfonamide nitrogen atom to provide compound 6. For example, 5 can becontacted with excess diazomethane (generated, for example, using1-methyl-3-nitro-1-nitrosoguanidine and sodium hydroxide) to afford 6where R⁶ is methyl. Other conventional alkylation procedures andreagents may also be employed to prepare various compounds of thisinvention.

[0214] In another preferred embodiment, compounds of this invention maybe prepared by displacement of a leaving group as shown in Scheme 2:

[0215] where HetAr and X are as defined herein; A′ is heteroaryl,substituted heteroaryl, heterocyclic or substituted heterocycliccontaining two nitrogen atoms in the heteroaryl or heterocyclic ring;and L¹ is a leaving group, such as chloro, bromo, iodo, sulfonate esterand the like.

[0216] Typically, this reaction is conducted by combining approximatelystoichiometric equivalents of 7 and 8 in a suitable inert diluent suchas water, dimethylsulfoxide (DMSO) and the like, with an excess of asuitable base such as sodium bicarbonate, sodium hydroxide, etc. toscavenge the acid generated by the reaction. The reaction is preferablyconducted at from about 25° C. to about 100° C. until reactioncompletion which typically occurs within 1 to about 24 hours. Thisreaction is further described in U.S. Pat. No. 3,598,859, which isincorporated herein by reference in its entirety. Upon reactioncompletion, the product 9 is recovered by conventional methods includingprecipitation, chromatography, filtration and the like.

[0217] In still another alternative embodiment, compounds of thisinvention can be prepared by reductive amination of a suitable2-oxocarboxylic acid ester, 10, such as a pyruvate ester, as shown inScheme 3:

[0218] where A′, HetAr and X are as defined herein.

[0219] Generally, this reaction is conducted by combining equimolaramounts of 10 and 11 in an inert diluent such as methanol, ethanol andthe like under conditions which provide for imine formation (not shown).The imine formed is then reduced under conventional conditions by asuitable reducing agent such as sodium cyanoborohydride, H₂/palladium oncarbon and the like to form the product 12. In a particularly preferredembodiment, the reducing agent is H₂/palladium on carbon which isincorporated into the initial reaction medium thereby permitting iminereduction in situ in a one pot procedure to provide 12. The reaction ispreferably conducted at from about 20° C. to about 80° C. at a pressureof from 1 to 10 atmospheres until reaction completion which typicallyoccurs within 1 to about 24 hours. Upon reaction completion, the product12 is recovered by conventional methods including chromatography,filtration and the like.

[0220] Alternatively, certain compounds of this invention can beprepared via a rhodium-catalyzed insertion reaction as shown in Scheme4:

[0221] where A″ is heteroaryl or substituted heteroaryl containing twonitrogen atoms in the heteroaryl ring, and HetAr and X (preferablyalkoxy) are as defined herein. Typically, this reaction is conductedusing rhodium acetate dimer, Rh₂(OAc)₄, in an inert diluent such astoluene at a temperature ranging from about 25° C. to about 80° C. forabout 1 to 12 hours to afford 15. This reaction is described further inB. R. Henke et. al., J. Med. Chem. 1998, 41, 5020-5036 and referencescited therein.

[0222] Similarly, certain compounds of this invention can be prepared bythe copper-catalyzed coupling reaction shown in Scheme 5:

[0223] where A″ is as defined herein, X³ is halogen, such as chloro,bromo or iodo (preferably iodo), and HetAr and X (preferably alkoxy) areas defined herein. Typically, this reaction is conducted using copperiodide (CuI) and potassium carbonate in an inert diluent such asN,N-dimethyl acetamide (DMA) at a temperature ranging from about 60° C.to about 120° C. for about 12 to 36 hours to afford 15. This reaction isdescribed further in D. Ma et. al., J. Am. Chem. Soc. 1998, 120,12459-12467 and references cited therein.

[0224] For ease of synthesis, the compounds of this invention aretypically prepared as an ester, i.e., where X is an alkoxy orsubstituted alkoxy group and the like. If desired, the ester group canbe hydrolysed using conventional conditions and reagents to provide thecorresponding carboxylic acid. Typically, this reaction is conducted bytreating the ester with at least one equivalent of an alkali metalhydroxide, such as lithium, sodium or potassium hydroxide, in an inertdiluent, such as methanol or mixtures of methanol and water, at atemperature ranging about 0° C. to about 24° C. for about 1 to about 12hours. Alternatively, benzyl esters may be removed by hydrogenolysisusing a palladium catalyst, such as palladium on carbon, and tert-butylesters can be removed using formic acid to afford the correspondingcarboxylic acid.

[0225] As will be apparent to those skilled in the art, other functionalgroups present on any of the substituents of the compounds of formulasI-II can be readily modified or derivatized either before or after theabove-described synthetic reactions using well-known syntheticprocedures. For example, a nitro group present on a substituent of acompound of Formula I-II or an intermediate thereof may be readilyreduced by hydrogenation in the presence of a palladium catalyst, suchas palladium on carbon, to provide the corresponding amino group. Thisreaction is typically conducted at a temperature of from about 20° C. toabout 50° C. for about 6 to about 24 hours in an inert diluent, such asmethanol.

[0226] Similarly, a pyridyl group can be hydrogenated in the presence ofa platinum catalyst, such as platinum oxide, in an acidic diluent toprovide the corresponding piperidinyl analogue. Generally, this reactionis conducted by treating the pyridine compound with hydrogen at apressure ranging from about 20 psi to about 60 psi, preferably about 40psi, in the presence of the catalyst at a temperature of about 20° C. toabout 50° C. for about 2 to about 24 hours in an acidic diluent, such asa mixture of methanol and aqueous hydrochloric acid.

[0227] Additionally, when the HetAr substituent of a compound of FormulaI-II or an intermediate thereof contains a primary or secondary aminogroup, such amino groups can be further derivatized either before orafter the above coupling reactions to provide, by way of example,amides, sulfonamides, ureas, thioureas, carbamates, secondary ortertiary amines and the like. Compounds having a primary amino group onthe HetAr substituent may be prepared, for example, by reduction of thecorresponding nitro compound as described above.

[0228] By way of illustration, a compound of Formula I-II or anintermediate thereof having a substituent containing a primary orsecondary amino group, such as where HetAr is a(4-aminopyridin-2-yl)methyl group, can be readily N-acylated usingconventional acylating reagents and conditions to provide thecorresponding amide. This acylation reaction is typically conducted bytreating the amino compound with at least one equivalent, preferablyabout 1.1 to about 1.2 equivalents, of a carboxylic acid in the presenceof a coupling reagent such as a carbodiimide, BOP reagent(benzotriazol-1-yloxy-tris(dimethylamino)-phosphoniumhexafluorophosphonate) and the like, in an inert diluent, such asdichloromethane, chloroform, acetonitrile, tetrahydrofuran,N,N-dimethylformamide and the like, at a temperature ranging from about0° C. to about 37° C. for about 4 to about 24 hours. Preferably, apromoter, such as N-hydroxysuccinimide, 1-hydroxy-benzotriazole and thelike, is used to facilitate the acylation reaction. Examples ofcarboxylic acids suitable for use in this reaction include, but are notlimited to, N-tert-butyloxycarbonylglycine,N-tert-butyloxycarbonyl-L-phenylalanine,N-tert-butyloxycarbonyl-L-aspartic acid benzyl ester, benzoic acid,N-tert-butyloxycarbonylisonipecotic acid, N-methylisonipecotic acid,N-tert-butyloxycarbonylnipecotic acid,N-tert-butyloxycarbonyl-L-tetrahydroisoquinoline-3-carboxylic acid,N-(toluene-4-sulfonyl)-L-proline and the like.

[0229] Alternatively, a compound of Formula I-II or an intermediatethereof containing a primary or secondary amino group can be N-acylatedusing an acyl halide or a carboxylic acid anhydride to form thecorresponding amide. This reaction is typically conducted by contactingthe amino compound with at least one equivalent, preferably about 1.1 toabout 1.2 equivalents, of the acyl halide or carboxylic acid anhydridein an inert diluent, such as dichloromethane, at a temperature rangingfrom about −70° C. to about 40° C. for about 1 to about 24 hours. Ifdesired, an acylation catalyst such as 4-(N,N-dimethylamino)pyridine maybe used to promote the acylation reaction. The acylation reaction ispreferably conducted in the presence of a suitablebase to scavenge theacid generated during the reaction. Suitable bases include, by way ofexample, tertiary amines, such as triethylamine, diisopropylethylamine,N-methylmorpholine and the like. Alternatively, the reaction can beconducted under Schotten-Baumann-type conditions using aqueous alkali,such as sodium hydroxide and the like.

[0230] Examples of acyl halides and carboxylic acid anhydrides suitablefor use in this reaction include, but are not limited to,2-methylpropionyl chloride, trimethylacetyl chloride, phenylacetylchloride, benzoyl chloride, 2-bromobenzoyl chloride, 2-methylbenzoylchloride, 2-trifluoro-methylbenzoyl chloride, isonicotinoyl chloride,nicotinoyl chloride, picolinoyl chloride, acetic anhydride, succinicanhydride, and the like. Carbamyl chlorides, such asN,N-dimethylcarbamyl chloride, N,N-diethylcarbamyl chloride and thelike, can also be used in this reaction to provide ureas. Similarly,dicarbonates, such as di-tert-butyl dicarbonate, may be employed toprovide carbamates.

[0231] In a similar manner, a compound of Formula I-II or anintermediate thereof containing a primary or secondary amino group maybe N-sulfonated to form a sulfonamide using a sulfonyl halide or asulfonic acid anhydride. Sulfonyl halides and sulfonic acid anhydridessuitable for use in this reaction include, but are not limited to,methanesulfonyl chloride, chloromethane-sulfonyl chloride,p-toluenesulfonyl chloride, trifluoromethanesulfonic anhydride, and thelike. Similarly, sulfamoyl chlorides, such as dimethylsulfamoylchloride, can be used to provide sulfamides (e.g., >N—SO₂—N<).

[0232] Additionally, a primary and secondary amino group present on asubstituent of a compound of Formula I-II or an intermediate thereof canbe reacted with an isocyanate or a thioisocyanate to give a urea orthiourea, respectively. This reaction is typically conducted bycontacting the amino compound with at least one equivalent, preferablyabout 1.1 to about 1.2 equivalents, of the isocyanate or thioisocyanatein an inert diluent, such as toluene and the like, at a temperatureranging from about 24° C. to about 37° C. for about 12 to about 24hours. The isocyanates and thioisocyanates used in this reaction arecommercially available or can be prepared from commercially availablecompounds using well-known synthetic procedures. For example,isocyanates and thioisocyanates are readily prepared by reacting theappropriate amine with phosgene or thiophosgene. Examples of isocyanatesand thioisocyanates suitable for use in this reaction include, but arenot limited to, ethyl isocyanate, n-propyl isocyanate, 4-cyanophenylisocyanate, 3-methoxyphenyl isocyanate, 2-phenylethyl isocyanate, methylthioisocyanate, ethyl thioisocyanate, 2-phenylethyl thioisocyanate,3-phenylpropyl thioisocyanate, 3-(N,N-diethylamino)propylthioisocyanate, phenyl thioisocyanate, benzyl thioisocyanate, 3-pyridylthioisocyanate, fluorescein isothiocyanate (isomer I) and the like.

[0233] Furthermore, when a compound of Formula I-II or an intermediatethereof contains a primary or secondary amino group, the amino group canbe reductively alkylated using aldehydes or ketones to form a secondaryor tertiary amino group. This reaction is typically conducted bycontacting the amino compound with at least one equivalent, preferablyabout 1.1 to about 1.5 equivalents, of an aldehyde or ketone and atleast one equivalent based on the amino compound of a metal hydridereducing agent, such as sodium cyanoborohydride, in an inert diluent,such as methanol, tetrahydrofuran, mixtures thereof and the like, at atemperature ranging from about 0° C. to about 50° C. for about 1 toabout 72 hours. Aldehydes and ketones suitable for use in this reactioninclude, by way of example, benzaldehyde, 4-chlorobenzaldehyde,valeraldehyde and the like.

[0234] In a similar manner, when a compound of Formula I-II or anintermediate thereof has a substituent containing a hydroxyl group, thehydroxyl group can be further modified or derivatized either before orafter the above coupling reactions to provide, by way of example,ethers, carbamates and the like. Compounds having a hydroxyl group onthe HetAr substituent, for example, can be prepared using an amino acidderivative derived from typtophan and the like in the above-describedreactions.

[0235] By way of example, a compound of Formula I-II or an intermediatethereof having a substituent containing a hydroxyl group, such as whereHetAr is a (5-hydroxypyridin-2-yl)methyl group, can be readilyO-alkylated to form ethers. This O-alkylation reaction is typicallyconducted by contacting the hydroxy compound with a suitable alkali oralkaline earth metal base, such as potassium carbonate, in an inertdiluent, such as acetone, 2-butanone and the like, to form the alkali oralkaline earth metal salt of the hydroxyl group. This salt is generallynot isolated, but is reacted in situ with at least one equivalent of analkyl or substituted alkyl halide or sulfonate, such as an alkylchloride, bromide, iodide, mesylate or tosylate, to afford the ether.Generally, this reaction is conducted at a temperature ranging fromabout 60° C. to about 150° C. for about 24 to about 72 hours.Preferably, a catalytic amount of sodium or potassium iodide is added tothe reaction mixture when an alkyl chloride or bromide is employed inthe reaction.

[0236] Examples of alkyl or substituted alkyl halides and sulfonatessuitable for use in this reaction include, but are not limited to,tert-butyl bromoacetate, N-tert-butyl chloroacetamide,1-bromoethylbenzene, ethyl α-bromophenylacetate,2-(N-ethyl-N-phenylamino)ethyl chloride, 2-(N,N-ethylamino)ethylchloride, 2-(N,N-diisopropylamino)ethyl chloride,2-(N,N-dibenzylamino)ethyl chloride, 3-(N,N-ethylamino)propyl chloride,3-(N-benzyl-N-methylamino)propyl chloride, N-(2-chloroethyl)morpholine,2-(hexamethyleneimino)ethyl chloride, 3-(N-methylpiperazine)propylchloride, 1-(3-chlorophenyl)-4-(3-chloropropyl)piperazine,2-(4-hydroxy-4-phenylpiperidine)ethyl chloride,N-tert-butyloxycarbonyl-3-piperidinemethyl tosylate, and the like.

[0237] Alternatively, a hydroxyl group present on a substituent of acompound of Formula I-II or an intermediate thereof can be O-alkylatingusing the Mitsunobu reaction. In this reaction, an alcohol, such as3-(N,N-dimethylamino)-1-propanol and the like, is reacted with about 1.0to about 1.3 equivalents of triphenylphosphine and about 1.0 to about1.3 equivalents of diethyl azodicarboxylate in an inert diluent, such astetrahydrofuran, at a temperature ranging from about −10° C. to about 5°C. for about 0.25 to about 1 hour. About 1.0 to about 1.3 equivalents ofa hydroxy compound, such as N-tert-butyltyrosine methyl ester, is thenadded and the reaction mixture is stirred at a temperature of about 0°C. to about 30° C. for about 2 to about 48 hours to provide theO-alkylated product.

[0238] In a similar manner, a compound of Formula I-II or anintermediate thereof containing an aryl or heteroaryl hydroxy group canbe reacted with an aryl iodide to provide a diaryl ether or a-heteroaryl-O-aryl ether. Generally, this reaction is conducted byforming the alkali metal salt of the hydroxyl group using a suitablebase, such as sodium hydride, in an inert diluent such as xylenes at atemperature of about −25° C. to about 10° C. The salt is then treatedwith about 1.1 to about 1.5 equivalents of cuprous bromide dimethylsulfide complex at a temperature ranging from about 10° C. to about 30°C. for about 0.5 to about 2.0 hours, followed by about 1.1 to about 1.5equivalents of an aryl iodide, such as sodium 2-iodobenzoate and thelike. The reaction is then heated to about 70° C. to about 150° C. forabout 2 to about 24 hours to provide the diaryl ether or the-heteroaryl-O-aryl ether.

[0239] Additionally, a hydroxy-containing compound can also be readilyderivatized to form a carbamate. In one method for preparing suchcarbamates, a hydroxy compound of Formula I-II or an intermediatethereof is contacted with about 1.0 to about 1.2 equivalents of4-nitrophenyl chloroformate in an inert diluent, such asdichloromethane, at a temperature ranging from about −25° C. to about 0°C. for about 0.5 to about 2.0 hours. Treatment of the resultingcarbonate with an excess, preferably about 2 to about 5 equivalents, ofa trialkylamine, such as triethylamine, for about 0.5 to 2 hours,followed by about 1.0 to about 1.5 equivalents of a primary or secondaryamine provides the carbamate. Examples of amines suitable for use inthis reaction include, but are not limited to, piperazine,1-methylpiperazine, 1-acetylpiperazine, morpholine, thiomorpholine,pyrrolidine, piperidine, N,N-dimethylamine and the like.

[0240] Alternatively, in another method for preparing carbamates, ahydroxy-containing compound is contacted with about 1.0 to about 1.5equivalents of a carbamyl chloride in an inert diluent, such asdichloromethane, at a temperature ranging from about 25° C. to about 70°C. for about 2 to about 72 hours. Typically, this reaction is conductedin the presence of a suitable base to scavenge the acid generated duringthe reaction. Suitable bases include, by way of example, tertiaryamines, such as triethylamine, diisopropylethylamine, N-methylmorpholineand the like. Additionally, at least one equivalent (based on thehydroxy compound) of 4-(N,N-dimethylamino)pyridine is preferably addedto the reaction mixture to facilitate the reaction. Examples of carbamylchlorides suitable for use in this reaction include, by way of example,dimethylcarbamyl chloride, diethylcarbamyl chloride and the like.

[0241] Likewise, when a compound of Formula I-II or an intermediatethereof contains a primary or secondary hydroxyl group, such hydroxylgroups can be readily converted into a leaving group and displaced toform, for example, amines, sulfides and fluorides. Generally, when achiral compound is employed in these reactions, the stereochemistry atthe carbon atom attached to the derivatized hydroxyl group is typicallyinverted.

[0242] These reactions are typically conducted by first converting thehydroxyl group into a leaving group, such as a tosylate, by treatment ofthe hydroxy compound with at least one equivalent of a sulfonyl halide,such as p-toluenesulfonyl chloride and the like, in pyridine. Thisreaction is generally conducted at a temperature of from about 0° C. toabout 70° C. for about 1 to about 48 hours. The resulting tosylate canthen be readily displaced with sodium azide, for example, by contactingthe tosylate with at least one equivalent of sodium azide in an inertdiluent, such as a mixture of N,N-dimethylformamide and water, at atemperature ranging from about 0° C. to about 37° C. for about 1 toabout 12 hours to provide the corresponding azido compound. The azidogroup can then be reduced by, for example, hydrogenation using apalladium on carbon catalyst to provide the amino (—NH₂) compound.

[0243] Similarly, a tosylate group can be readily displaced by a thiolto form a sulfide. This reaction is typically conducted by contactingthe tosylate with at least one equivalent of a thiol, such asthiophenol, in the presence of a suitable base, such as1,8-diazabicyclo[5.4.0]undec-7-ene (DBU), in an inert diluent, such asN,N-dimethylformamide, at a temperature of from about 0° C. to about 37°C. for about 1 to about 12 hours to provide the sulfide. Additionally,treatment of a tosylate with morpholinosulfur trifluoride in an inertdiluent, such as dichloromethane, at a temperature ranging from about 0°C. to about 37° C. for about 12 to about 24 hours affords thecorresponding fluoro compound.

[0244] Furthermore, a compound of Formula I-II or an intermediatethereof having a substituent containing an iodoheteroaryl group, forexample, when HetAr is a (4-iodopyridyl)methyl group, can be readilyconverted either before or after the above coupling reactions into a-heteroaryl-aryl compound. Typically, this reaction is conducted bytreating the iodoheteroaryl compound with about 1.1 to about 2equivalents of an arylzinc iodide, such as 2-(methoxycarbonyl)phenylzinciodide, in the presence of a palladium catalyst, such as palladiumtetra(triphenylphosphine), in an inert diluent, such as tetrahydrofuran,at a temperature ranging from about 24° C. to about 30° C. untilreaction completion. This reaction is further described, for example, inRieke, J. Org. Chem. 1991, 56, 1445. Additional methods for preparing-heteroaryl-aryl derivatives are disclosed in International PublicationNumber WO 98/53817, published Dec. 3, 1998, the disclosure of which isincorporated herein by reference in its entirety. The -heteroaryl-arylcompound can also be prepared by coupling (4-iodopyridyl)methyl groupwith arylboronic acid under Suzuki coupling reaction conditions wellknown in the art.

[0245] In some cases, the compounds of Formula I-II or intermediatesthereof may contain substituents having one or more sulfur atoms. Whenpresent, such sulfur atoms can be oxidized either before or after theabove coupling reactions to provide a sulfoxide or sulfone compoundusing conventional reagents and reaction conditions. Suitable reagentsfor oxidizing a sulfide compound to a sulfoxide include, by way ofexample, hydrogen peroxide, 3-chloroperoxy-benzoic acid (MCPBA), sodiumperiodate and the like. The oxidation reaction is typically conducted bycontacting the sulfide compound with about 0.95 to about 1.1 equivalentsof the oxidizing reagent in an inert diluent, such as dichloromethane,at a temperature ranging from about −50° C. to about 75° C. for about 1to about 24 hours. The resulting sulfoxide can then be further oxidizedto the corresponding sulfone by contacting the sulfoxide with at leastone additional equivalent of an oxidizing reagent, such as hydrogenperoxide, MCPBA, potassium permanganate and the like. Alternatively, thesulfone can be prepared directly by contacting the sulfide with at leasttwo equivalents, and preferably an excess, of the oxidizing reagent.Such reactions are described further in March, “Advanced OrganicChemistry”, 4th Ed., pp. 1201-1202, Wiley Publisher, 1992.

[0246] Other procedures and reaction conditions for preparing thecompounds of this invention are described in the examples set forthbelow. Additionally, other procedures for preparing compounds useful incertain aspects of this invention are disclosed in U.S. application Ser.Nos. 09/489,377 and 09/489,378, filed on Jan. 21, 2000, entitled“Compounds Which Inhibit Leucocyte Adhesion Mediated by VLA-4” (AttorneyDocket Nos. 002010-517 and 002010-525); the disclosures of which areincorporated herein by reference in their entirety.

[0247] Pharmaceutical Formulations

[0248] When employed as pharmaceuticals, the compounds of this inventionare usually administered in the form of pharmaceutical compositions.These compounds can be administered by a variety of routes includingoral, rectal, transdermal, subcutaneous, intravenous, intramuscular, andintranasal. These compounds are effective as both injectable and oralcompositions. Such compositions are prepared in a manner well known inthe pharmaceutical art and comprise at least one active compound.

[0249] This invention also includes pharmaceutical compositions whichcontain, as the active ingredient, one or more of the compounds ofFormula I-II above associated with pharmaceutically acceptable carriers.In making the compositions of this invention, the active ingredient isusually mixed with an excipient, diluted by an excipient or enclosedwithin such a carrier which can be in the form of a capsule, sachet,paper or other container. The excipient employed is typically anexcipient suitable for administration to human subjects or othermammals. When the excipient serves as a diluent, it can be a solid,semi-solid, or liquid material, which acts as a vehicle, carrier ormedium for the active ingredient. Thus, the compositions can be in theform of tablets, pills, powders, lozenges, sachets, cachets, elixirs,suspensions, emulsions, solutions, syrups, aerosols (as a solid or in aliquid medium), ointments containing, for example, up to 10% by weightof the active compound, soft and hard gelatin capsules, suppositories,sterile injectable solutions, and sterile packaged powders.

[0250] In preparing a formulation, it may be necessary to mill theactive compound to provide the appropriate particle size prior tocombining with the other ingredients. If the active compound issubstantially insoluble, it ordinarily is milled to a particle size ofless than 200 mesh. If the active compound is substantially watersoluble, the particle size is normally adjusted by milling to provide asubstantially uniform distribution in the formulation, e.g. about 40mesh.

[0251] Some examples of suitable excipients include lactose, dextrose,sucrose, sorbitol, mannitol, starches, gum acacia, calcium phosphate,alginates, tragacanth, gelatin, calcium silicate, microcrystallinecellulose, polyvinylpyrrolidone, cellulose, water, syrup, and methylcellulose. The formulations can additionally include: lubricating agentssuch as talc, magnesium stearate, and mineral oil; wetting agents;emulsifying and suspending agents; preserving agents such as methyl- andpropylhydroxy-benzoates; sweetening agents; and flavoring agents. Thecompositions of the invention can be formulated so as to provide quick,sustained or delayed release of the active ingredient afteradministration to the patient by employing procedures known in the art.

[0252] The compositions are preferably formulated in a unit dosage form,each dosage containing from about 5 to about 100 mg, more usually about10 to about 30 mg, of the active ingredient. The term “unit dosageforms” refers to physically discrete units suitable as unitary dosagesfor human subjects and other mammals, each unit containing apredetermined quantity of active material calculated to produce thedesired therapeutic effect, in association with a suitablepharmaceutical excipient.

[0253] The active compound is effective over a wide dosage range and isgenerally administered in a pharmaceutically effective amount. It, willbe understood, however, that the amount of the compound actuallyadministered will be determined by a physician, in the light of therelevant circumstances, including the condition to be treated, thechosen route of administration, the actual compound administered, theage, weight, and response of the individual patient, the severity of thepatient's symptoms, and the like.

[0254] For preparing solid compositions such as tablets, the principalactive ingredient is mixed with a pharmaceutical excipient to form asolid preformulation composition containing a homogeneous mixture of acompound of the present invention. When referring to thesepreformulation compositions as homogeneous, it is meant that the activeingredient is dispersed evenly throughout the composition so that thecomposition may be readily subdivided into equally effective unit dosageforms such as tablets, pills and capsules. This solid preformulation isthen subdivided into unit dosage forms of the type described abovecontaining from, for example, 0.1 to about 500 mg of the activeingredient of the present invention.

[0255] The tablets or pills of the present invention may be coated orotherwise compounded to provide a dosage form affording the advantage ofprolonged action. For example, the tablet or pill can comprise an innerdosage and an outer dosage component, the latter being in the form of anenvelope over the former. The two components can be separated by anenteric layer which serves to resist disintegration in the stomach andpermit the inner component to pass intact into the duodenum or to bedelayed in release. A variety of materials can be used for such entericlayers or coatings, such materials including a number of polymeric acidsand mixtures of polymeric acids with such materials as shellac, cetylalcohol, and cellulose acetate.

[0256] The liquid forms in which the novel compositions of the presentinvention may be incorporated for administration orally or by injectioninclude aqueous solutions suitably flavored syrups, aqueous or oilsuspensions, and flavored emulsions with edible oils such as cottonseedoil, sesame oil, coconut oil, or peanut oil, as well as elixirs andsimilar pharmaceutical vehicles.

[0257] Compositions for inhalation or insufflation include solutions andsuspensions in pharmaceutically acceptable, aqueous or organic solvents,or mixtures thereof, and powders. The liquid or solid compositions maycontain suitable pharmaceutically acceptable excipients as describedsupra. Preferably the compositions are administered by the oral or nasalrespiratory route for local or systemic effect. Compositions inpreferably pharmaceutically acceptable solvents may be nebulized by useof inert gases. Nebulized solutions may be breathed directly from thenebulizing device or the nebulizing device may be attached to a facemasks tent, or intermittent positive pressure breathing machine.Solution, suspension, or powder compositions may be administered,preferably orally or nasally, from devices which deliver the formulationin an appropriate manner.

[0258] The following formulation examples illustrate the pharmaceuticalcompositions of the present invention.

Formulation Example 1

[0259] Hard gelatin capsules containing the following ingredients areprepared: Quantity Ingredient (mg/capsule) Active Ingredient 30.0 Starch305.0 Magnesium stearate 5.0

[0260] The above ingredients are mixed and filled into hard gelatincapsules in 340 mg quantities.

Formulation Example 2

[0261] A tablet formula is prepared using the ingredients below:Quantity Ingredient (mg/tablet) Active Ingredient 25.0 Cellulose,microcrystalline 200.0 Colloidal silicon dioxide 10.0 Stearic acid 5.0

[0262] The components are blended and compressed to form tablets, eachweighing 240 mg.

Formulation Example 3

[0263] A dry powder inhaler formulation is prepared containing thefollowing components: Ingredient Weight % Active Ingredient 5 Lactose 95

[0264] The active mixture is mixed with the lactose and the mixture isadded to a dry powder inhaling appliance.

Formulation Example 4

[0265] Tablets, each containing 30 mg of active ingredient, are preparedas follows: Quantity Ingredient (mg/tablet) Active Ingredient 30.0 mgStarch 45.0 mg Microcrystalline cellulose 35.0 mg Polyvinylpyrrolidone 4.0 mg (as 10% solution in water) Sodium carboxymethyl starch  4.5 mgMagnesium stearate  0.5 mg Talc  1.0 mg Total  120 mg

[0266] The active ingredient, starch and cellulose are passed through aNo. 20 mesh U.S. sieve and mixed thoroughly. The solution ofpolyvinyl-pyrrolidone is mixed with the resultant powders, which arethen passed through a 16 mesh U.S. sieve. The granules so produced aredried at 50° to 60° C. and passed through a 16 mesh U.S. sieve. Thesodium carboxymethyl starch, magnesium stearate, and talc, previouslypassed through a No. 30 mesh U.S. sieve, are then added to the granuleswhich, after mixing, are compressed on a tablet machine to yield tabletseach weighing 150 mg.

Formulation Example 5

[0267] Capsules, each containing 40 mg of medicament are made asfollows: Quantity Ingredient (mg/capsule) Active Ingredient  40.0 mgStarch 109.0 mg Magnesium stearate  1.0 mg Total 150.0 mg

[0268] The active ingredient, cellulose, starch, an magnesium stearateare blended, passed through a No. 20 mesh U.S. sieve, and filled intohard gelatin capsules in 150 mg quantities.

Formulation Example 6

[0269] Suppositories, each containing 25 mg of active ingredient aremade as follows: Ingredient Amount Active Ingredient   25 mg Saturatedfatty acid glycerides to 2,000 mg

[0270] The active ingredient is passed through a No. 60 mesh U.S. sieveand suspended in the saturated fatty acid glycerides previously meltedusing the minimum heat necessary. The mixture is then poured into asuppository mold of nominal 2.0 g capacity and allowed to cool.

Formulation Example 7

[0271] Suspensions, each containing 50 mg of medicament per 5.0 ml doseare made as follows: Ingredient Amount Active Ingredient 50.0 mg Xanthangum  4.0 mg Sodium carboxymethyl cellulose (11%) 50.0 mgMicrocrystalline cellulose (89%) Sucrose 1.75 g Sodium benzoate 10.0 mgFlavor and Color q.v. Purified water to  5.0 ml

[0272] The medicament, sucrose and xanthan gum are blended, passedthrough a No. 10 mesh U.S. sieve, and then mixed with a previously madesolution of the microcrystalline cellulose and sodium carboxymethylcellulose in water. The sodium benzoate, flavor, and color are dilutedwith some of the water and added with stirring. Sufficient water is thenadded to produce the required volume.

Formulation Example 8

[0273] Quantity Ingredient (mg/capsule) Active Ingredient  15.0 mgStarch 407.0 mg Magnesium stearate  3.0 mg Total 425.0 mg

[0274] The active ingredient, cellulose, starch, and magnesium stearateare blended, passed through a No. 20 mesh U.S. sieve, and filled intohard gelatin capsules in 560 mg quantities.

Formulation Example 9

[0275] An intravenous formulation may be prepared as follows: IngredientQuantity Active Ingredient 250.0 mg Isotonic saline  1000 mL

Formulation Example 10

[0276] A topical formulation may be prepared as follows: IngredientOuantity Active Ingredient 1-10 g Emulsifying Wax 30 g Liquid Paraffin20 g White Soft Paraffin to 100 g

[0277] The white soft paraffin is heated until molten. The liquidparaffin and emulsifying wax are incorporated and stirred untildissolved. The active ingredient is added and stirring is continueduntil dispersed. The mixture is then cooled until solid.

[0278] Another preferred formulation employed in the methods of thepresent invention employs transdermal delivery devices (“patches”). Suchtransdermal patches may be used to provide continuous or discontinuousinfusion of the compounds of the present invention in controlledamounts. The construction and use of transdermal patches for thedelivery of pharmaceutical agents is well known in the art. See, e.g.,U.S. Pat. No. 5,023,252, issued Jun. 11, 1991, herein incorporated byreference. Such patches may be constructed for continuous, pulsatile, oron demand delivery of pharmaceutical agents.

[0279] Direct or indirect placement techniques may be used when it isdesirable or necessary to introduce the pharmaceutical composition tothe brain. Direct techniques usually involve placement of a drugdelivery catheter into the host's ventricular system to bypass theblood-brain barrier. One such implantable delivery system used for thetransport of biological factors to specific anatomical regions of thebody is described in U.S. Pat. No. 5,011,472 which is hereinincorporated by reference.

[0280] Indirect techniques, which are generally preferred, usuallyinvolve formulating the compositions to provide for drug latentiation bythe conversion of hydrophilic drugs into lipid-soluble drugs.Latentiation is generally achieved through blocking of the hydroxy,carbonyl, sulfate, and primary amine groups present on the drug torender the drug more lipid soluble and amenable to transportation acrossthe blood-brain barrier. Alternatively, the delivery of hydrophilicdrugs may be enhanced by intra-arterial infusion of hypertonic solutionswhich can transiently open the blood-brain barrier.

[0281] Utility

[0282] The compounds of this invention can be employed to bind VLA-4α₄β₁ integrin) in biological samples, i.e., the compounds bind VLA-4with an IC₅₀ of 15 μM or less in a competitive binding assay asdescribed herein. Accordingly, these compounds have utility in, forexample, assaying such samples for VLA-4. In such assays, the compoundscan be bound to a solid support and the VLA-4 sample added thereto. Theamount of VLA-4 in the sample can be determined by conventional methodssuch as use of a sandwich ELISA assay. Alternatively, labeled VLA-4 canbe used in a competitive assay to measure for the presence of VLA-4 inthe sample. Other suitable assays are well known in the art.

[0283] In addition, certain of the compounds of this invention inhibit,in vivo, adhesion of leukocytes to endothelial cells mediated by VLA-4by competitive binding to VLA-4. Accordingly, the compounds of thisinvention can be used in the treatment of diseases mediated by VLA-4 orleucocyte adhesion. Such diseases include inflammatory diseases inmammalian patients such as asthma, Alzheimer's disease, atherosclerosis,AIDS dementia, diabetes (including acute juvenile onset diabetes),inflammatory bowel disease (including ulcerative colitis and Crohn'sdisease), multiple sclerosis, rheumatoid arthritis, tissuetransplantation, tumor metastasis, meningitis, encephalitis, stroke, andother cerebral traumas, nephritis, retinitis, atopic dermatitis,psoriasis, myocardial ischemia and acute leukocyte-mediated lung injurysuch as that which occurs in adult respiratory distress syndrome.

[0284] The biological activity of the compounds identified above may beassayed in a variety of systems. For example, a compound can beimmobilized on a solid surface and adhesion of cells expressing VLA-4can be measured. Using such formats, large numbers of compounds can bescreened. Cells suitable for this assay include any leukocytes known toexpress VLA-4 such as T cells, B cells, monocytes, eosinophils, andbasophils. A number of leukocyte cell lines can also be used, examplesinclude Jurkat and U937.

[0285] The test compounds can also be tested for the ability tocompetitively inhibit binding between VLA-4 and VCAM-1, or between VLA-4and a labeled compound known to bind VLA-4 such as a compound of thisinvention or antibodies to VLA-4. In these assays, the VCAM-1 can beimmobilized on a solid surface. VCAM-1 may also be expressed as arecombinant fusion protein having an Ig tail (e.g., IgG) so that bindingto VLA-4 may be detected in an immunoassay. Alternatively, VCAM-1expressing cells, such as activated endothelial cells or VCAM-1transfected fibroblasts can be used. For assays to measure the abilityto block adhesion to brain endothelial cells, the assays described inInternational Patent Application Publication No. WO 91/05038 areparticularly preferred. This application is incorporated herein byreference in its entirety.

[0286] Many assay formats employ labeled assay components. The labelingsystems can be in a variety of forms. The label may be coupled directlyor indirectly to the desired component of the assay according to methodswell known in the art. A wide variety of labels may be used. Thecomponent may be labelled by any one of several methods. The most commonmethod of detection is the use of autoradiography with ³H, ¹²⁵I, ³⁵S,¹⁴C, or ³²P labeled compounds or the like. Non-radioactive labelsinclude ligands which bind to labeled antibodies, fluorophores,chemiluminescent agents, enzymes and antibodies which can serve asspecific binding pair members for a labeled ligand. The choice of labeldepends on sensitivity required, ease of conjugation with the compound,stability requirements, and available instrumentation.

[0287] Appropriate in vivo models for demonstrating efficacy in treatinginflammatory responses include EAE (experimental autoimmuneencephalomyelitis) in mice, rats, guinea pigs or primates, as well asother inflammatory models dependent upon α4 integrins.

[0288] Compounds having the desired biological activity may be modifiedas necessary to provide desired properties such as improvedpharmacological properties (e.g., in vivo stability, bio-availability),or the ability to be detected in diagnostic applications. Stability canbe assayed in a variety of ways such as by measuring the half-life ofthe proteins during incubation with peptidases or human plasma or serum.A number of such protein stability assays have been described (see,e.g., Verhoef et al., Eur. J. Drug Metab. Pharmacokinet., 1990,15(2):83-93).

[0289] For diagnostic purposes, a wide variety of labels may be linkedto the compounds, which may provide, directly or indirectly, adetectable signal. Thus, the compounds of the subject invention may bemodified in a variety of ways for a variety of end purposes while stillretaining biological activity. In addition, various reactive sites maybe introduced at the terminus for linking to particles, solidsubstrates, macromolecules, or the like.

[0290] Labeled compounds can be used in a variety of in vivo or in vitroapplications. A wide variety of labels may be employed, such asradionuclides (e.g., gamma-emitting radioisotopes such as technetium-99or indium-111), fluorescers (e.g., fluorescein), enzymes, enzymesubstrates, enzyme cofactors, enzyme inhibitors, chemiluminescentcompounds, bioluminescent compounds, and the like. Those of ordinaryskill in the art will know of other suitable labels for binding to thecomplexes, or will be able to ascertain such using routineexperimentation. The binding of these labels is achieved using standardtechniques common to those of ordinary skill in the art.

[0291] In vitro uses include diagnostic applications such as monitoringinflammatory responses by detecting the presence of leukocytesexpressing VLA-4. The compounds of this invention can also be used forisolating or labeling such cells. In addition, as mentioned above, thecompounds of the invention can be used to assay for potential inhibitorsof VLA-4/VCAM-1 interactions.

[0292] For in vivo diagnostic imaging to identify, e.g., sites ofinflammation, radioisotopes are typically used in accordance with wellknown techniques. The radioisotopes may be bound to the peptide eitherdirectly or indirectly using intermediate functional groups. Forinstance, chelating agents such as diethylenetriaminepentacetic acid(DTPA) and ethylenediaminetetraacetic acid (EDTA) and similar moleculeshave been used to bind proteins to metallic ion radioisotopes.

[0293] The complexes can also be labeled with a paramagnetic isotope forpurposes of in vivo diagnosis, as in magnetic resonance imaging (MRI) orelectron spin resonance (ESR), both of which are well known. In general,any conventional method for visualizing diagnostic imaging can be used.Usually gamma- and positron-emitting radioisotopes are used for cameraimaging and paramagnetic isotopes are used for MRI. Thus, the compoundscan be used to monitor the course of amelioration of an inflammatoryresponse in an individual. By measuring the increase or decrease inlymphocytes expressing VLA-4 it is possible to determine whether aparticular therapeutic regimen aimed at ameliorating the disease iseffective.

[0294] The pharmaceutical compositions of the present invention can beused to block or inhibit cellular adhesion associated with a number ofdiseases and disorders. For instance, a number of inflammatory disordersare associated with integrins or leukocytes. Treatable disordersinclude, e.g., transplantation rejection (e.g., allograft rejection),Alzheimer's disease, atherosclerosis, AIDS dementia, diabetes (includingacute juvenile onset diabetes), retinitis, cancer metastases, rheumatoidarthritis, acute leukocyte-mediated lung injury (e.g., adult respiratorydistress syndrome), asthma, nephritis, and acute and chronicinflammation, including atopic dermatitis, psoriasis, myocardialischemia, and inflammatory bowel disease (including Crohn's disease andulcerative colitis). In preferred embodiments the pharmaceuticalcompositions are used to treat inflammatory brain disorders, such asmultiple sclerosis (MS), viral meningitis and encephalitis.

[0295] Inflammatory bowel disease is a collective term for two similardiseases referred to as Crohn's disease and ulcerative colitis. Crohn'sdisease is an idiopathic, chronic ulceroconstrictive inflammatorydisease characterized by sharply delimited and typically transmuralinvolvement of all layers of the bowel wall by a granulomatousinflammatory reaction. Any segment of the gastrointestinal tract, fromthe mouth to the anus, may be involved, although the disease mostcommonly affects the terminal ileum and/or colon. Ulcerative colitis isan inflammatory response limited largely to the colonic mucosa andsubmucosa. Lymphocytes and macrophages are numerous in lesions ofinflammatory bowel disease and may contribute to inflammatory injury.

[0296] Asthma is a disease characterized by increased responsiveness ofthe tracheobronchial tree to various stimuli potentiating paroxysmalconstriction of the bronchial airways. The stimuli cause release ofvarious mediators of inflammation from IgE-coated mast cells includinghistamine, eosinophilic and neutrophilic chemotactic factors,leukotrines, prostaglandin and platelet activating factor. Release ofthese factors recruits basophils, eosinophils and neutrophils, whichcause inflammatory injury.

[0297] Atherosclerosis is a disease of arteries (e.g., coronary,carotid, aorta and iliac). The basic lesion, the atheroma, consists of araised focal plaque within the intima, having a core of lipid and acovering fibrous cap. Atheromas compromise arterial blood flow andweaken affected arteries. Myocardial and cerebral infarcts are a majorconsequence of this disease. Macrophages and leukocytes are recruited toatheromas and contribute to inflammatory injury.

[0298] Rheumatoid arthritis is a chronic, relapsing inflammatory diseasethat primarily causes impairment and destruction of joints. Rheumatoidarthritis usually first affects the small joints of the hands and feetbut then may involve the wrists, elbows, ankles and knees. The arthritisresults from interaction of synovial cells with leukocytes thatinfiltrate from the circulation into the synovial lining of the joints.See e.g., Paul, Immunology (3d ed., Raven Press, 1993).

[0299] Another indication for the compounds of this invention is intreatment of organ or graft rejection mediated by VLA-4. Over recentyears there has been a considerable improvement in the efficiency ofsurgical techniques for transplanting tissues and organs such as skin,kidney, liver, heart, lung, pancreas and bone marrow. Perhaps theprincipal outstanding problem is the lack of satisfactory agents forinducing immunotolerance in the recipient to the transplanted allograftor organ. When allogeneic cells or organs are transplanted into a host(i.e., the donor and donee are different individuals from the samespecies), the host immune system is likely to mount an immune responseto foreign antigens in the transplant (host-versus-graft disease)leading to destruction of the transplanted tissue. CD8⁺ cells, CD4 cellsand monocytes are all involved in the rejection of transplant tissues.Compounds of this invention which bind to alpha-4 integrin are useful,inter alia, to block alloantigen-induced immune responses in the doneethereby preventing such cells from participating in the destruction ofthe transplanted tissue or organ. See, e.g., Paul et al., TransplantInternational 9, 420-425 (1996); Georczynski et al., Immunology 87,573-580 (1996); Georcyznski et al., Transplant. Immunol. 3, 55-61(1995); Yang et al., Transplantation 60, 71-76 (1995); Anderson et al.,APMIS 102, 23-27 (1994).

[0300] A related use for compounds of this invention which bind to VLA-4is in modulating the immune response involved in “graft versus host”disease (GVHD). See e.g., Schlegel et al., J. Immunol. 155, 3856-3865(1995). GVHD is a potentially fatal disease that occurs whenimmunologically competent cells are transferred to an allogeneicrecipient. In this situation, the donor's immunocompetent cells mayattack tissues in the recipient. Tissues of the skin, gut epithelia andliver are frequent targets and may be destroyed during the course ofGVHD. The disease presents an especially severe problem when immunetissue is being transplanted, such as in bone marrow transplantation;but less severe GVHD has also been reported in other cases as well,including heart and liver transplants. The therapeutic agents of thepresent invention are used, inter alia, to block activation of the donorT-cells thereby interfering with their ability to lyse target cells inthe host.

[0301] A further use of the compounds of this invention is inhibitingtumor metastasis. Several tumor cells have been reported to expressVLA-4 and compounds which bind VLA-4 block adhesion of such cells toendothelial cells. Steinback et al., Urol. Res. 23, 175-83 (1995); Oroszet al., Int. J. Cancer 60, 867-71 (1995); Freedman et al., Leuk.Lymphoma 13, 47-52 (1994); Okahara et al., Cancer Res. 54, 3233-6(1994).

[0302] A further use of the compounds of this invention is in treatingmultiple sclerosis. Multiple sclerosis is a progressive neurologicalautoimmune disease that affects an estimated 250,000 to 350,000 peoplein the United States. Multiple sclerosis is thought to be the result ofa specific autoimmune reaction in which certain leukocytes attack andinitiate the destruction of myelin, the insulating sheath covering nervefibers. In an animal model for multiple sclerosis, murine monoclonalantibodies directed against VLA-4 have been shown to block the adhesionof leukocytes to the endothelium, and thus prevent inflammation of thecentral nervous system and subsequent paralysis in the animals¹⁶.

[0303] Pharmaceutical compositions of the invention are suitable for usein a variety of drug delivery systems. Suitable formulations for use inthe present invention are found in Remington's Pharmaceutical Sciences,Mace Publishing Company, Philadelphia, Pa., 17th ed. (1985).

[0304] In order to enhance serum half-life, the compounds may beencapsulated, introduced into the lumen of liposomes, prepared as acolloid, or other conventional techniques may be employed which providean extended serum half-life of the compounds. A variety of methods areavailable for preparing liposomes, as described in, e.g., Szoka, et al.,U.S. Pat. Nos. 4,235,871, 4,501,728 and 4,837,028 each of which isincorporated herein by reference.

[0305] The amount administered to the patient will vary depending uponwhat is being administered, the purpose of the administration, such asprophylaxis or therapy, the state of the patient, the manner ofadministration, and the like. In therapeutic applications, compositionsare administered to a patient already suffering from a disease in anamount sufficient to cure or at least partially arrest the symptoms ofthe disease and its complications. An amount adequate to accomplish thisis defined as “therapeutically effective dose.” Amounts effective forthis use will depend on the disease condition being treated as well asby the judgment of the attending clinician depending upon factors suchas the severity of the inflammation, the age, weight and generalcondition of the patient, and the like.

[0306] The compositions administered to a patient are in the form ofpharmaceutical compositions described above. These compositions may besterilized by conventional sterilization techniques, or may be sterilefiltered. The resulting aqueous solutions may be packaged for use as is,or lyophilized, the lyophilized preparation being combined with asterile aqueous carrier prior to administration. The pH of the compoundpreparations typically will be between 3 and 11, more preferably from 5to 9 and most preferably from 7 to 8. It will be understood that use ofcertain of the foregoing excipients, carriers, or stabilizers willresult in the formation of pharmaceutical salts.

[0307] The therapeutic dosage of the compounds of the present inventionwill vary according to, for example, the particular use for which thetreatment is made, the manner of administration of the compound, thehealth and condition of the patient, and the judgment of the prescribingphysician. For example, for intravenous administration, the dose willtypically be in the range of about 20 μg to about 500 μg per kilogrambody weight, preferably about 100 μg to about 300 μg per kilogram bodyweight. Suitable dosage ranges for intranasal administration aregenerally about 0.1 pg to 1 mg per kilogram body weight. Effective dosescan be extrapolated from dose-response curves derived from in vitro oranimal model test systems.

[0308] Compounds of this invention are also capable of binding orantagonizing the actions of α₆β_(i), α₉β₁, α₄β₇, α_(d)β₂, α_(e)β₇integrins (although α₄β₁ and α₉β₁ are preferred in this invention).Accordingly, compounds of this invention are also useful for preventingor reversing the symptoms, disorders or diseases induced by the bindingof these integrins to their respective ligands.

[0309] For example, International Publication Number WO 98/53817,published Dec. 3, 1998 (the disclosure of which is incorporated hereinby reference in its entirety) and references cited therein describedisorders mediated by α₄β₇. This reference also describes an assay fordetermining antagonism of α₄β₇ dependent binding to VCAM-Ig fusionprotein.

[0310] Additionally, compounds that bind α_(d)β₂ and α_(e)β₇ integrinsare particularly useful for the treatment of asthma and related lungdiseases. See, for example, M. H. Grayson et al., J. Exp. Med. 1998,188(11) 2187-2191. Compounds that bind α_(e)β₇ integrin are also usefulfor the treatment of systemic lupus erythematosus (see, for example, M.Pang et al., Arthritis Rheum. 1998, 41(8), 1456-1463); Crohn's disease,ulcerative colitis and infammatory bowel disease (IBD) (see, forexample, D. Elewaut et al., Scand J. Gastroenterol 1998, 33(7) 743-748);Sjogren's syndrome (see, for example, U. Kroneld et al., Scand J.Gastroenterol 1998, 27(3), 215-218); and rheumatoid arthritis (see, forexample, Scand J. Gastroenterol 1996, 44(3), 293-298). And compoundsthat bind α₆β₁ may be useful in preventing fertilization (see, forexample, H. Chen et al., Chem. Biol. 1999, 6, 1-10).

[0311] Certain of the compounds within the generic formulas describedherein are also useful as synthetic intermediates for other compounds ofthis invention as illustrated in the examples herein.

[0312] The following synthetic and biological examples are offered toillustrate this invention and are not to be construed in any way aslimiting the scope of this invention. Unless otherwise stated, alltemperatures are in degrees Celsius.

EXAMPLES

[0313] In the examples below, the following abbreviations have thefollowing meanings. If an abbreviation is not defined, it has itsgenerally accepted meaning. aq or aq. = aqueous AcOH = acetic acid bd =broad doublet bm = broad multiplet bs = broad singlet Bn = benzyl Boc =N-tert-butoxylcarbonyl Boc₂O = di-tert-butyl dicarbonate BOP =benzotriazol-1-yloxy- tris(dimethylamino)phosphonium hexafluorophosphateCbz = carbobenzyloxy CHCl₃ = chloroform CH₂Cl₂ = dichloromethane (COCl)₂= oxalyl chloride d = doublet dd = doublet of doublets dt = doublet oftriplets DBU = 1,8-diazabicyclo[5.4.0]undec-7-ene DCC =1,3-dicyclohexylcarbodiimide DMAP = 4-N,N-dimethylaminopyridine DME =ethylene glycol dimethyl ether DMF = N,N-dimethylformamide DMSO =dimethylsulfoxide EDC = 1-(3-dimethylaminopropyl)-3- ethylcarbodiimidehydrochloride Et₃N = triethylamine Et₂O = diethyl ether EtOAc = ethylacetate EtOH = ethanol eq or eq. = equivalent Fmoc =N-(9-fluorenylmethoxycarbonyl) FmocONSu =N-(9-fluorenylmethoxycarbonyl)- succinimide g = grams h = hour H₂O =water HBr = hydrobromic acid HCl = hydrochloric acid HOBT =1-hydroxybenzotriazole hydrate hr = hour K₂CO₃ = potassium carbonate L =liter m = multiplet MeOH = methanol mg = milligram MgSO₄ = magnesiumsulfate mL = milliliter mm = millimeter mM = millimolar mmol = millimolmp = melting point N = normal NaCl = sodium chloride Na₂CO₃ = sodiumcarbonate NaHCO₃ = sodium bicarbonate NaOEt = sodium ethoxide NaOH =sodium hydroxide NH₄Cl = ammonium chloride NMM = N-methylmorpholine Phe= L-phenylalanine Pro = L-proline psi = pounds per square inch PtO₂ =platinum oxide q = quartet quint. = quintet rt = room temperature s =singlet sat = saturated t = triplet t-BuOH = tert-butanol TFA =trifluoroacetic acid THF = tetrahydrofuran TLC or tlc = thin layerchromatography Ts = tosyl TsCl = tosyl chloride TsOH = tosylate μL =microliter

[0314] The following Methods may be used to prepare the compounds ofthis invention.

Method A 5-Trifluoromethylsulfonyloxy-2-methylpyridine PreparationProcedure

[0315] The procedure of Tilley et al. J. Org. Chem. 1998, 5, 3(2),386-390, was used to convert 5-hydroxy-2-methylpyridine into5-trifluoromethyl-sulfonyloxy-2-methylpyridine.

Method B Heteroaryl-aryl Preparation Procedure—Suzuki Coupling Procedure

[0316] A mixture of a heteroaryl halide or heteroaryl triflate (1.0eq.), an arylboronic acid (1.1 eq.), CsCO₃ (1.5 eq.), Pd(PPh₃)₄ (0.03eq.) And anhydrous DMF was stirred under nitrogen at 100° C. for 48 hand then the DMF was evaporated. The residue was partitioned betweenethyl acetate and half-saturated aqueous sodium chloride and then theethyl acetate extracts were treated with magnesium sulfate, filtered andevaporated. The residue was purified by flash chromatography on silicagel using ethyl acetate/hexanes to give the heteroaryl-aryl product.

Method C 3-Heteroarylpyruvic Acid Ethyl Ester Preparation Procedure

[0317] To a stirred −78° C. solution of a methylheteroaryl compound (1eq.) in tetrahydrofuran was added dropwise 1.6 M n-butyllithium inhexanes (1.25 eq.). The resulting dark red solution was warmed to 0° C.and then re-cooled to −78° C. A solution of diethyl oxalate (1.25 eq.)in THF was added dropwise. The resulting mixture was warmed to 22° C.,stirred for 18 h diluted with aqueous HCl, and the extracted withmethylene chloride. The methylene chloride extracts were treated withmagnesium sulfate, filtered, and evaporated. The residue was purified byflash chromatography on silica gel column using ethyl acetate/hexanes togive a 3-heteroarylpyruvic acid ethyl ester.

Method D 3-Heteroarylpyruvic Acid Ethyl Ester Oxime PreparationProcedure

[0318] The procedure of Negi et al. Synthesis, 1996, 8, 991, was used toconvert a 3-heteroarylpyruvic acid ethyl ester into a3-heteroarylpyruvic ethyl ester oxime.

Method E 3-Heteroarylalanine Acid Ethyl Ester Preparation Procedure

[0319] The procedure of Keck et al. Syn. Commun., 1979, 281-286 was usedto convert a 3-heteroarylpyruvic acid ethyl ester into aD,L-3-heteroarylalanine ethyl ester.

Method F 3-Heteroaryldehydroalanine Methyl Ester Preparation Procedure

[0320] The reaction of N-benzyloxycarbonyldehydroalanine methyl esterwith an aryl bromide according to the procedure of Carlstrom and Frejd.J. Org. Chem., 1991, 56, 1289-1293, was used to prepare a3-heteroaryldehydroalanine methyl ester.

Method G 3-Heteroarylalanine Methyl Ester Oxime Preparation Procedure

[0321] The procedure of Keck et al. Syn. Commun., 1979, 281-286 was usedto convert a 3-heteroaryldehydroalanine methyl ester into aD,L-3-heteroarylalanine methyl ester.

Method H Methyl Ester Preparation Procedure

[0322] Amino acid methyl esters can be prepared using the method ofBrenner and Huber Helv. Chim. Acta 1953, 36, 1109.

Method I BOP Coupling Procedure

[0323] The desired dipeptide ester was prepared by the reaction of acarboxylic acid (1 equivalent) with the appropriate amino acid ester oramino acid ester hydrochloride (1 equivalent),benzotriazol-1-yloxy-tris(dimethylamino)phos-phonium hexafluorophosphate[BOP] (2.0 equivalent), triethylamine (1.1 equivalent), and DMF. Thereaction mixture was stirred at room temperature overnight. The crudeproduct is purified flash chromatography to afford the dipeptide ester.

Method J Hydrogenation Procedure I

[0324] Hydrogenation was performed using 10% palladium on carbon (10% byweight) in methanol at 30 psi overnight. The mixture was filteredthrough a pad of Celite and the filtrate concentrated to yield thedesired compound.

Method K Hydrolysis Procedure I

[0325] To a chilled (0° C.) THF/H₂O solution (2:1, 5-10 mL) of theappropriate ester was added LiOH (or NaOH) (0.95 equivalents). Thetemperature was maintained at 0° C. and the reaction was complete in 1-3hours. The reaction mixture was extracted with ethyl acetate and theaqueous phase was lyophilized resulting in the desired carboxylate salt.

Method L Ester Hydrolysis Procedure II

[0326] To a chilled (0° C.) THF/H₂O solution (2:1, 5-10 mL) of theappropriate ester was added LiOH (1.1 equivalents). The temperature wasmaintained at 0° C. and the reaction was complete in 1-3 hours. Thereaction mixture was concentrated and the residue was taken up into H₂Oand the pH adjusted to 2-3 with aqueous HCl. The product was extractedwith ethyl acetate and the combined organic phase was washed with brine,dried over MgSO₄, filtered and concentrated to yield the desired acid.

Method M Ester Hydrolysis Procedure III

[0327] The appropriate ester was dissolved in dioxane/H₂O (1:1) and 0.9equivalents of 0.5 N NaOH was added. The reaction was stirred for 3-16hours and then concentrated. The resulting residue was dissolved in H₂Oand extracted with ethyl acetate. The aqueous phase was lyophilized toyield the desired carboxylate sodium salt.

Method N BOC Removal Procedure

[0328] Anhydrous hydrochloride (HCl) gas was bubbled through amethanolic solution of the appropriate Boc-amino acid ester at 0° C. for15 minutes and the reaction mixture was stirred for three hours. Thesolution was concentrated to a syrup and dissolved in Et₂O andreconcentrated. This procedure was repeated and the resulting solid wasplaced under high vacuum overnight.

Method O Tert-Butyl Ester Hydrolysis Procedure I

[0329] The tert-butyl ester was dissolved in CH₂Cl₂ and treated withTFA. The reaction was complete in 1-3 hr at which time the reactionmixture was concentrated and the residue dissolved in H₂O andlyophilized to yield the desired acid.

Method P EDC Coupling Procedure I

[0330] To a CH₂Cl₂ solution (5-20 mL) of a carboxylic acid (1equivalent), the appropriate amino acid ester hydrochloride (1equivalent), N-methylmorpholine (1.1-2.2 equivalents) and1-hydroxybenzotriazole (2 equivalents) were mixed, placed in an ice bathand 1-(3-dimethylaminopropyl)-3-ethyl carbodiimide (1.1 equivalents)added. The reaction was allowed to rise to room temperature and stirredovernight. The reaction mixture was poured into H₂O and the organicphase was washed with sat. NaHCO₃, brine, dried (NgSO₄ or Na₂SO₄),filtered and concentrated. The crude product was purified by columnchromatography.

Method Q EDC Coupling Procedure II

[0331] To a DMF solution (5-20 mL) of a carboxylic acid (1 equivalent),the appropriated amino acid ester hydrochloride (1 equivalent), Et₃N(1.1 equivalents) and 1-hydroxybenzotriazole (2 equivalents) were mixed,placed in an ice bath and 1-(3-dimethylaminopropyl)-3-ethyl carbodiimide(1.1 equivalents) added. The reaction was allowed to rise to roomtemperature and stirred overnight. The reaction mixture was partitionedbetween EtOAc and H₂O and the organic phase washed with 0.2 N citricacid, H₂O, sat. NaHCO₃, brine, dried (MgSO₄ or Na₂SO₄), filtered andconcentrated. The crude product was purified by column chromatography orpreparative TLC.

Method R Tert-Butyl Ester Hydrolysis Procedure II

[0332] The tert-butyl ester was dissolved in CH₂Cl₂ (5 mL) and treatedwith TFA (5 mL). The reaction was complete in 1-3 hours at which timethe reaction mixture was concentrated and the residue dissolved in H₂Oand concentrated. The residue was redissolved in H₂O and lyophilized toyield the desired product.

Method S Carbamate Formation Procedure I

[0333] Into a reaction vial were combined 15.2 mmol, 1.0 eq. of thestarting hydroxy compound (typically a tyrosine derivative) and 1.86 g(15.2 mmol, 1.0 eq) DMAP. Methylene chloride (50 mL), triethylamine(2.12 mL, 1.54 g, 15.2 mmol, 1.0 eq), and dimethylcarbamyl chloride(1.68 mL, 1.96 g, 18.2 mmol, 1.2 eq) were then added. The vial wascapped tightly, and the reaction solution swirled to obtain ahomogeneous solution. The reaction solution was then heated to 40° C.After 48 h, TLC of the resulting colorless solution indicated completeconversion. The work-up of the reaction solution was as follows: 50 mLEtOAc and 50 mL hexanes was added to the reaction mixture, and theresulting mixture was washed with 0.5 M citric acid (3×50 mL), water(2×50 mL), 10% K₂CO₃ (2×50 mL), and sat. NaCl (1×50 mL); dried withMgSO₄, filtered and evaporated to afford the desired compound.

Method T Carbamate Formation Procedure II

[0334] Into a reaction vial were combined 84.34 mmol (1.0 eq) of thestarting hydroxy compound (typically a tyrosine derivative) and 17.0 g(84.34 mmol, 1.0 eq) 4-nitrophenyl chloroformate. Methylene chloride(700 mL) was added and the vial was capped with a septum. A nitrogenline was attached and the vial was immersed in a 4:1 water/ethanol dryice slurry with stirring to cool to −15° C. Triethylamine (29.38 mL,21.33 g, 210.81 mmol, 2.5 eq) was added over five minutes with stirringand the stirring was continued at −10 to −15° C. for 1 h. N-Methylpiperazine (9.35 mL, 8.45 g, 84.34 mmol, 1.0 eq) was added over threeminutes with stirring and stirring was continued overnight while warmingto room temperature. The reaction mixture was diluted with 700 mLhexanes and the resulting mixture was washed repeatedly with 10% K₂CO₃,until no yellow color (from 4-nitrophenol) is observed in the aqueouslayer. The mixture was then washed with sat. NaCl, dried over anhydrousMgSO₄, filtered and evaporated. The residue was dissolved in 500 mL ofethanol and evaporated to remove triethylamine. The residue was againdissolved in 500 mL of ethanol and evaporated to remove triethylamine.The residue was then dissolved in 400 mL of ethanol and 600 mL of waterwas added with stirring to precipitate a solid or oil. If an oil ifformed, the oil is stirred vigorously to induce it to solidify. Thesolid is then isolated by filtration. Dissolution, precipitation, andfiltration are repeated once and the resulting solid is rinsed withwater to remove traces of yellow color. The solid is then subjected tohigh vacuum until the mass remains constant thereby affording thedesired carbamyloxy compound.

Method U Preparation of 5-Iodo-4(3H)-pyrimidinone

[0335] The procedure of Sakamoto et. al. (Chem. Pharm. Bull. 1986,34(7), 2719-2724) was used to convert 4(3H)-pyrimidinone into5-iodo-4(3H)-pyrimidinone, which was of sufficient purity for conversionto 4-chloro-5-iodopyrimidine.

Method V Preparation of 4-Chloro-5-iodopyrimidine

[0336] 5-Iodo-4(3H)-pyrimidinone (1 eq.) was suspended in toluene towhich was added POCl₃ (2.0 eq.). The reaction mixture was heated toreflux for 3 hours, and then cooled and concentrated. The residue wassuspended in water, adjusted to pH=7 by addition of 4N sodium hydroxide,and extracted with ethyl acetate. The organic extracts were washed withbrine, dried (MgSQ₄), filtered and stripped to give a red oil. The crudeproduct was dissolved in methanol and silica gel was added. Followingconcentration, the coated silica gel was loaded onto a plug of silicagel and elution with ethyl acetate/hexanes yielded the title compound.

Method W Preparation of 4-N-Alkylamino-5-bromo-2-chloropyrimidine

[0337] A methanol solution of 5-bromo-2,4-dichloropyrimidine (1.0 eq),the alkylamine (1.05 eq, typicallyL-4-(N,N-dimethylcarbamyloxy)phenylalanine tert-butyl ester), andN,N-diisoproylethylamine (5.0 eq) was heated to 40° C. for 16 hours. Thereaction mixture was then concentrated, and the residue was taken up inethyl acetate. The organic portion was washed with 0.2 N citric acid,water, saturated NaHCO₃, brine, dried (MgSO₄), filtered andconcentrated. The crude material was purified by silica gelchromatography using ethyl acetate/hexanes to afford the desiredcompound.

Method X Preparation of 4-N-Alkylamino-5-bromo-2-N-alkylaminopyrimidine

[0338] An isopropanol solution of the4-N-alkylamino-5-bromo-2-chloropyrimidine (1.0 eq) and an alkylamine(5.0 eq) was heated in sealed tube at 130° C. for 3-5 hours. Thereaction mixture was then cooled and washed with 0.2 N citric acid,water, saturated NaHCO₃, brine, dried (MgSO₄), filtered andconcentrated. The crude material was purified by silica gelchromatography using ethyl acetate/hexanes to afford the desiredcompound.

Method Y 4-N-Alkylamino-5-bromo-2-N-alkylaminopyrimidine Suzuki CouplingProcedure

[0339] To an ethyleneglycol dimethyl ether solution oftetrakis(triphenylphosphine)palladium (0.04 eq) was added an4-N-alkylamino-5-bromo-2-N-alkylaminopyrimidine (1.0 eq.). Afterstirring for approximately ten minutes, the boronic acid or ester (1.2eq) and 2M Na₂CO₃ (2.0 eq) was added, and the reaction flask wasevacuated and then flushed with nitrogen gas. The reaction was heated atreflux for three to four hours. The reaction mixture was then cooled anddiluted with water and ethyl acetate, and the organic phase was washedwith 0.2 N citric acid, water, saturated NaHCO₃, brine, dried (MgSO₄),filtered and concentrated. The residue was purified by either silica gelcolumn or preparative thin layer chromatography using ethylacetate/hexanes to afford the desired product.

Method Z Preparation of Dimethyl 2-Alkylmalonate

[0340] To a suspension of sodium hydride 60% dispersion in mineral oil(1.1 eq) in anhydrous THF was added slowly with stirring dimethylmalonate (1.1 eq), causing the evolution of gas. To the resultingsolution was added a bromoalkane, iodoalkane, ortrifluoromethanesulfonyloxyalkane (1.0 eq), and the mixture was heatedto 50° C. for 48 h, at which point TLC indicated consumption of thebromoalkane, iodoalkane, or trifluoromethanesulfonyloxyalkane. Themixture was diluted with diethyl ether and washed with 70% saturatedsodium chloride. The organic extracts were treated with anhydrousmagnesium sulfate, filtered, and evaporated to afford a dimethyl2-alkylmalonate of sufficient purity for immediate conversion to a5-alkyl-4,6-dihydroxypyrimidine.

Method AA Preparation of Diethyl 2-Alkylidenylmalonate

[0341] Procedure B (p. 2759) of Houve and Winberg (J. Org. Chem. 1980,45(14), 2754-2763) was employed to react diethyl malonate with a ketoneor an aldehyde to afford a diethyl 2-alkylidenylmalonate of sufficientpurity for immediate conversion to a diethyl 2-alkylmalonate.

Method BB Preparation of Diethyl 2-Alkylmalonate

[0342] A diethyl 2-alkylidenylmalonate and an equal mass 10% palladiumon carbon were suspended in ethanol. The mixture was shaken under 55 psihydrogen gas for 24 h, at which point TLC indicated consumption of thediethyl 2-alkylidenylmalonate. The mixture was filtered through Celiteand evaporated to afford a diethyl 2-alkylmalonate of sufficient purityfor immediate conversion to a 5-alkyl-4,6-dihydroxypyrimidine.

Method CC Preparation of 5-Alkyl-4,6-dihydroxypyrimidine

[0343] To a diethyl 2-alkylmalonate or a dimethyl 2-alkylmalonate (1.0eq) was added formamidine acetate (1.0 eq) and 25% sodium methoxide inmethanol (3.3 eq). The resulting slurry was stirred vigorously andheated to 60° C. for 4 h, and then allowed to cool. The slurry wasdiluted with water, and acidified to pH=2 by addition of HCl. Theresulting precipitate was collected by filtration, washed with water,and dried under vacuum, to afford a 5-alkyl-4,6-dihydroxypyrimidine ofsufficient purity for immediate conversion to a5-alkyl-4,6-dichloropyrimidine.

Method DD Preparation of 5-Alkoxy-4-hydroxypyrimidine

[0344] The method (p. 308) of Anderson et al. (Org. Proc. Res. Devel.1997, 1, 300-310) was employed to react a methyl alkoxyacetate, sodiummethoxide, ethyl formate, and formamidine acetate to afford a5-alkoxy-4-hydroxypyrimidine of sufficient purity for immediateconversion to a 5-alkoxy-4-chloropyrimidine.

Method EE Preparation of 5-Alkyl-4,6-dichloropyrimidine or5-Alkoxy-4-chloropyrimidine

[0345] To a 5-alkyl-4,6-dihydroxypyrimidine or a5-alkoxy-4-hydroxypyrimidine (1.0 eq) were added phosphorus oxychloride(15.0 eq) and N,N-dimethylaniline (1.0 eq), and the mixture was heatedto 100° C. for 3 h, and then allowed to cool. The resulting solution waspoured onto ice, and the mixture was extracted with dichloromethane. Theorganic extracts were treated with anhydrous magnesium sulfate,filtered, and evaporated to afford a 5-alkyl-4,6-dichloropyrimidine or a5-alkoxy-4-chloropyrimidine of sufficient purity for immediateconversion to a 5-alkyl-4-N-alkylamino-6-chloropyrimidine or a5-alkoxy-4-N-alkylaminopyrimidine.

Method FF Preparation of 5-Alkyl-4-N-alkylamino-6-chloropyrimidine or5-Alkoxy-4-N-alkylaminopyrimidine

[0346] To a solution of a 5-alkyl-4,6-dichloropyrimidine or a5-alkoxy-4-chloropyrimidine (1.0 eq) in ethanol were added an alkylamine (1.2 eq, typically L-4-(N,N-dimethylcarbamyloxy)-phenylalaninetert-butyl ester) and diisopropylethylamine (2.0 eq). The mixture wassealed in a pressure tube and heated to 120° C. for 48 h, at which pointTLC indicated consumption of the 5-alkyl-4,6-dichloropyrimidine or the5-alkoxy-4-chloropyrimidine. The mixture was evaporated, and the residuewas partitioned between ethyl acetate and pH=4.5 citrate buffer. Theorganic extracts were washed with saturated sodium chloride, treatedwith anhydrous magnesium sulfate, filtered, and evaporated. The residuewas purified by chromatography on silica gel using ethyl acetate andhexanes to afford a pure 5-alkyl-4-N-alkylamino-6-chloropyrimidine or5-alkoxy-4-N-alkylaminopyrimidine.

Method GG Preparation of 5-Alkyl-4-N-alkylaminopyrimidine (Procedure I)

[0347] A suspension of 5-alkyl-4-N-alkylamino-6-chloropyrimidine (1.0eq), and an equal mass 10% palladium on carbon, and sodium bicarbonate(5.0 eq) in methanol was shaken under 55 psi hydrogen gas for 16 h, atwhich point TLC indicated consumption of the5-alkyl-4-N-alkylamino-6-chloropyrimidine. The mixture was filteredthrough Celite and evaporated to give a residue, which was partitionedbetween ethyl acetate and 70% saturated sodium chloride. The organicextracts were treated with anhydrous magnesium sulfate, filtered, andevaporated. The residue was purified by chromatography on silica gelusing ethyl acetate and hexanes to afford a pure5-alkyl-4-N-alkylaminopyrimidine.

Method HH Preparation of 5-Alkyl-4-N-alkylaminopyrimidine (Procedure II)

[0348] A suspension of 5-alkyl-4-N-alkylamino-6-chloropyrimidine (1.0eq), sodium acetate (10.0 eq), and zinc powder (20.0 eq) in a 9:1mixture of acetic acid and water was stirred vigorously at 40° C. for 72h, at which point TLC indicated partial consumption of the5-alkyl-4-N-alkylamino-6-chloropyrimidine. The supernatant solution wasdecanted from remaining zinc and evaporated. The residue was partitionedbetween ethyl acetate and saturated sodium bicarbonate, and the organicextracts were treated with anhydrous magnesium sulfate, filtered, andevaporated. The residue was purified by chromatography on silica gelusing ethyl acetate and hexanes to afford a pure5-alkyl-4-N-alkylaminopyrimidine.

Method II Preparation of 2,4-Dichloro-5-nitropyrimidine

[0349] 5-Nitrouracil was treated with phosphorus oxychloride andN,N-dimethylaniline, according to the procedure of Whittaker (J. Chem.Soc. 1951, 1565), to give 2,4-dichloro-5-nitropyrimidine as an orangeoil, which was used without distillation immediately in the next step.

Method JJ Preparation of Diethyl 2-(N,N-Dialkylamino)malonate

[0350] The appropriate amine (1.0 eq) was added to a 0° C. solution ofdiethyl bromomalonate (1.0 eq) and N,N-diisopropylethyl amine (1.1 eq)in ethanol. The mixture was stirred and allowed to warm roomtemperature. After 16 hours, the reaction mixture was concentrated andthe residue was suspended in ethyl acetate and sat. NaHCO₃. The organicportion was washed with sat NaHCO₃, brine, dried (MgSO₄) filtered andconcentrated to yield the diethyl 2-(N,N-dialkylamino)malonate, ofsufficient purity for immediate conversion to a5-(N,N-dialkylamino)-4,6-dihydroxypyrimidine.

Method KK Preparation of 5-(N,N-Dialkylamino)-4,6-dihydroxypyrimidine

[0351] A suspension of a diethyl 2-(N,N-dialkylamino)malonate (1.0 eq),formamidine acetate (1.10 eq.) and 25% sodium methoxide in methanol (3.3eq) was heated to 65° C. for 3.5 hours. The reaction mixture was cooledand diluted with water. The mixture was acidified to pH=4.5 by additionof dilute HCl. The resulting precipitate was collected by filtration,washed with water, and dried under vacuum to afford a5-(N,N-dialkylamino)-4,6-dihydroxy-pyrimidine of sufficient purity forimmediate conversion to a 5-(N,N-dialkylamino)-4,6-dichloropyrimidine.Alternatively, the acidified solution was evaporated to give a solidresidue, which was extracted with boiling ethanol. The ethanol extractswere filtered and concentrated to give a residue, which wasrecrystallized from isopropyl alcohol to afford a5-(N,N-dialkylamino)-4,6-dihydroxypyrimidine of sufficient purity forimmediate conversion to a 5-(N,N-dialkylamino)-4,6-dichloropyrimidine.

Method LL Preparation of 5-(N,N-Dialkylamino)-4,6-dichloropyrimidine

[0352] A 5-(N,N-dialkylamino)-4,6-dihydroxypyrimidine (1.0 eq) wassuspended in POCl₃ (15.0 eq), and the mixture was heated to reflux for16 hours. Then the mixture was cooled and carefully poured into asuspension of ethyl ether and aqueous K₂CO₃. The organic portion waswashed with brine, dried (MgSO₄), filtered and concentrated to yield a5-(N,N-dialkylamino)-4,6-dichloro-pyrimidine of sufficient purity forimmediate reaction with alkylamines.

Method MM Preparation of4-(N-Alkylamino)-5-(N,N-dialkylamino)-6-chloropyrimidine

[0353] A 5-(N,N-dialkylamino)4,6-dichloropyrimidine (1.0 eq),L-4-(N,N-dimethylcarbamyloxy)phenylalanine tert-butyl ester (1.5 eq) andN,N-diisopropyl ethylamine (1.5 eq) were dissolved in ethanol and heatedto 120° C. in a sealed tube for 72 h. The cooled reaction mixture wasconcentrated, and the residue dissolved in ethyl acetate. The ethylacetate solution was washed with 0.2 N citric acid, water, saturatedNaHCO₃, brine, dried (MgSO₄), filtered and concentrated. The residue waspurified by silica gel chromatography using ethyl acetate/hexanes toafford the 4-(N-alkylamino)-5-(N,N-dialkylamino)-6-chloropyrimidine.

Method NN Preparation of 4-(N-Alkylamino)-5-(N,N-dialkylamino)pyrimidine

[0354] A 4-(N-Alkylamino)-5-(N,N-dialkylamino)-6-chloropyrimidine (1.0eq), an equal mass of 10% palladium on carbon, and NaHCO₃ (5.0 eq) weresuspended in methanol. The reaction mixture was hydrogenated at 45 psihydrogen for 16 hours and then filtered through a pad of Celite. Thefiltrate was concentrated, and the residue was dissolved in ethylacetate. The ethyl acetate solution was washed with water, brine, dried(MgSO₄), filtered and concentrated to yield an oil. The oil was purifiedby column chromatography on silica gel using ethyl actate and hexanes toafford a pure 4-(N-alkylamino)-5-(N,N-dialkylamino)pyrimidine.

Method OO Bromopyrimidine Debromination Procedure

[0355] The bromopyrimidine was dissolved in isopropyl alcohol to whichwas added 10% palladium on carbon. The reaction was hydrogenated at 45psi hydrogen. Filtration and concentration of the filtrate yielded thedesired dehalogenated pyrimidine.

Method PP Preparation of 2-Isopropropoxypyrimidine

[0356] A 2-chloropyrimidine was dissolved in isopropyl alcohol to whichwas added diisopropylamine. The reaction was heated in a sealed tube forten days at 130° C. The cooled reaction mixture was concentrated, andthe product purified via silica gel column chromatography to yield the2-isopropoxypyrimidine.

Method QQ Preparation of 2-Amino-3-Chloropyrazine

[0357] A mixture of 2,3-dichloropyrazine (Lancaster) and ammoniumhydroxide was heated in a sealed tube at 100° C. for 24 h resulting in awhite precipitate. The precipitate was collected by filtration and driedunder vacuum to afford 2-amino-3-chloropyrazine of sufficient purity forimmediate conversion to 2-chloro-3-nitropyrazine.

Method RR Preparation of 2-Chloro-3-Nitropyrazine

[0358] The method (p. 1638) of Hartman et al. (J. Med. Chem. 1984,27(12), 1634-1639) was employed to convert 2-amino-3-chloropyrazine into2-chloro-3-nitropyrazine of sufficient purity for immediate use.

Method SS Preparation of 4-Alkylamino-2-dialkylamino-5-nitropyrimidine

[0359] A solution of 1.0 eq 4-alkylamino-2-chloro-5-nitropyrimidine and5.0 eq dialkylamine in THF was allowed to stand for 16 h. The mixturewas diluted with ethyl acetate and then washed with pH=4.5 citratebuffer and saturated sodium chloride. The organic extracts were treatedwith anhydrous magnesium sulfate, filtered, and evaporated to give aresidue, which was purified by chromatography on silica gel using ethylacetate and hexanes.

Method TT Preparation of 5-Hydroxypyridin-2-ylalanine

[0360] The methods of Schow et al. (J. Org. Chem. 1994 59:22,6850-6852), Ye et al (J. Org. Chem. 1995 60:8, 2640-2641) or Myers etal. (J. Org. Chem. 1996 61:22, 813-815) may be used to prepare the titlecompound.

Method UU Preparation of 2-Hydroxypyridin-5-ylalanine

[0361] The methods of Yamano et al. (Tetrahedron. 1992 48:8, 1457-1464)or Andrews et al. (J. Chem. Soc. Perkin Trans. 1995 11, 1335-1340) maybe used to prepare the title compound.

Example 1 Synthesis ofN-(5-(2,2,2-trifluoroethyl)pyrimidin-4-yl)-D,L-3-(5-(2,5-dimethoxyphenyl)-pyridin-2-yl)alanine

[0362] 5-Hydroxy-2-methylpyridine was converted via method A to give5-trifluoromethylsulfonyloxy-2-methylpyridine. This compound and2,6-dimethoxyphenylboronic acid were coupled via method B to give5-(2,6-dimethoxyphenyl)-2-methylpyridine which was then converted viasequential application of methods C,D and E to giveD,L-(5-(2,6-dimethoxy-phenyl)pyridin-2-yl)alanine ethyl ester.

[0363] 2,2,2-Trifluoroethyl (prepared accordingly to Gassman, et al. J.Org. Chem., 1984, 49(12), 2258-2273) was converted via sequentialapplication of methods Z, CC, and EE to give4,6-dichloro-5-(2,2,2-trifluoroethyl)pyrimidine.D,L-(5-(2,6-dimethoxyphenyl)pyridine-2-yl)alanine ethyl ester and4,6-dichloro-5-(2,2,2-trifluoroethyl)pyrimidine were coupled via methodFF and the product was transformed via sequential application of methodsGG and L to give the title compound.

Example 2 Synthesis ofN-(5-(2,2,2-trifluoroethyl)pyrimidin-4-yl)-D,L-3-(5-(2-methoxyphenyl)-pyridin-2-yl)alanine

[0364] 5-Hydroxy-2-methylpyridine was converted via method A to give5-trifluoromethylsulfonyloxy-2-methylpyridine. This compound and2-methoxyphenylboronic acid were coupled via method B to give5-(2-methoxyphenyl)-2-methylpyridine which was then converted viasequential application of methods C,D and E to giveD,L-(5-(2-methoxyphenyl)pyridin-2-yl)alanine ethyl ester.

[0365] 2,2,2-Trifluoroethyl (prepared accordingly to Gassman, et al. J.Org. Chem., 1984, 49(12), 2258-2273) was converted via sequentialapplication of methods Z, CC, and EE to give4,6-dichloro-5-(2,2,2-trifluoroethyl)pyrimidine.D,L-(5-(2-methoxyphenyl)pyridine-2-yl)alanine ethyl ester and4,6-dichloro-5-(2,2,2-trifluoroethyl)pyrimidine were coupled via methodFF and the product was transformed via sequential application of methodsGG and L to give the title compound.

Example 3 Synthesis ofN-(5-(2,2,2-trifluoroethyl)pyrimidin-4-yl)-D,L-3-(5-(N,N-dimethylaminocarbonyloxy)-pyridin-2-yl)alanine

[0366] 5-Hydroxypyridin-2-ylalanine, which can be prepared as perExample TT, is N and C protected via conventional methods to provide forethyl N-Boc 5-hydroxypyridin-2-ylalanine. This compound is convertedeither by methods S or T to provide for ethyl N-Boc5-(N,N-dimethylaminocarbonyloxypyridin-2-ylalanine. Conventionaldeprotection provides for5-(N,N-dimethylaminocarbonyloxypyridin-2-ylalanine.

[0367] Coupling of the amino of the5-(N,N-dimethylaminocarbonyloxypyridin-2-ylalanine with4,6-dichloro-5-(2,2,2-trifluoroethyl)pyrimidine as per Example 2 abovewill provide the title compound.

Example 4 Synthesis ofN-(5-(2,2,2-trifluoroethyl)pyrimidin-4-yl)-D,L-3-(2-(N,N-dimethylaminocarbonyloxy)-pyridin-5-yl)alanine

[0368] 2-Hydroxypyridin-5-ylalanine, which can be prepared as perExample UU, is N and C protected via conventional methods to provide forethyl N-Boc 2-hydroxypyridin-5-ylalanine. This compound is convertedeither by methods S or T to provide for ethyl N-Boc2-(N,N-dimethylaminocarbonyloxypyridin-5-ylalanine. Conventionaldeprotection provides for2-(N,N-dimethylaminocarbonyloxypyridin-5-ylalanine.

[0369] Coupling of the amino of the2-(N,N-dimethylaminocarbonyloxypyridin-5-ylalanine with4,6-dichloro-5-(2,2,2-trifluoroethyl)pyrimidine as per Example 2 abovewill provide the title compound.

Biological Examples Example A In Vitro Assay For Determining Binding ofCandidate Compounds to VLA-4

[0370] An in vitro assay was used to assess binding of candidatecompounds to α₄β₁ integrin. Compounds which bind in this assay can beused to assess VCAM-1 levels in biological samples by conventionalassays (e.g., competitive assays). This assay is sensitive to IC₅₀values as low as about 1 nM.

[0371] The activity of α₄β₁ integrin was measured by the interaction ofsoluble VCAM-1 with Jurkat cells (e.g., American Type Culture CollectionNos. TIB 152, TIB 153, and CRL 8163), a human T-cell line whichexpresses high levels of α₄β₁ integrin. VCAM-1 interacts with the cellsurface in an α₄β₁ integrin-dependent fashion (Yednock, et al. J. Biol.Chem., 1995, 270:28740).

[0372] Recombinant soluble VCAM-1 was expressed as a chimeric fusionprotein containing the seven extracellular domains of VCAM-1 on theN-terminus and the human IgG₁ heavy chain constant region on theC-terminus. The VCAM-1 fusion protein was made and purified by themanner described by Yednock, supra.

[0373] Jurkat cells were grown in RPMI 1640 supplemented with 10% fetalbovine serum, penicillin, streptomycin and glutamine as described byYednock, supra.

[0374] Jurkat cells were incubated with 1.5 mM MnCl₂ and 5 μg/mL 15/7antibody for 30 minutes on ice. Mn⁺² activates the receptor to enhanceligand binding, and 15/7 is a monoclonal antibody that recognizes anactivated/ligand occupied conformation of α₄β₁ integrin and locks themolecule into this conformation thereby stabilizing the VCAM-1/α₄β₁integrin interaction. Yednock, et al., supra. Antibodies similar to the15/7 antibody have been prepared by other investigators (Luque, et al,1996, J. Biol. Chem. 271:11067) and may be used in this assay.

[0375] Cells were then incubated for 30 minutes at room temperature withcandidate compounds, in various concentrations ranging from 66 μM to0.01 μM using a standard 5-point serial dilution. 15 μL solublerecombinant VCAM-1 fusion protein was then added to Jurkat cells andincubated for 30 minutes on ice. (Yednock et al., supra.).

[0376] Cells were then washed two times and resuspended in PE-conjugatedgoat F(ab′)₂ anti-mouse IgG Fc (Immunotech, Westbrook, Me.) at 1:200 andincubated on ice, in the dark, for 30 minutes. Cells were washed twiceand analyzed with a standard fluorescence activated cell sorter (“FACS”)analysis as described in Yednock, et al., supra.

[0377] Compounds having an IC₅₀ of less than about 15 μM possess bindingaffinity to α₄β₁.

[0378] When tested in this assay, each of the compound prepared in theabove examples has or is expected to have an IC₅₀ of 15 1μM or less (oris expected to be active in vivo).

Example B In vitro Saturation Assay For Determining Binding of CandidateCompounds to α₄β₁

[0379] The following describes an in vitro assay to determine the plasmalevels needed for a compound to be active in the Experimental AutoimmuneEncephalomyelitis (“EAE”) model, described in the next example, or inother in vivo models.

[0380] Log-growth Jurkat cells are washed and resuspended in normalanimal plasma containing 20 μg/ml of the 15/7 antibody (described in theabove example).

[0381] The Jurkat cells are diluted two-fold into either normal plasmasamples containing known candidate compound amounts in variousconcentrations ranging from 66 μM to 0.01 μM, using a standard 12 pointserial dilution for a standard curve, or into plasma samples obtainedfrom the peripheral blood of candidate compound-treated animals.

[0382] Cells are then incubated for 30 minutes at room temperature,washed twice with phosphate-buffered saline (“PBS”) containing 2% fetalbovine serum and 1 mM each of calcium chloride and magnesium chloride(assay medium) to remove unbound 15/7 antibody.

[0383] The cells are then exposed to phycoerythrin-conjugated goatF(ab′)₂ anti-mouse IgG Fc (Immunotech, Westbrook, Me.), which has beenadsorbed for any non-specific cross-reactivity by co-incubation with 5%serum from the animal species being studied, at 1:200 and incubated inthe dark at 4° C. for 30 minutes.

[0384] Cells are washed twice with assay medium and resuspended in thesame. They are then analyzed with a standard fluorescence activated cellsorter (“FACS”) analysis as described in Yednock et al. J. Biol. Chem.,1995, 270:28740.

[0385] The data is then graphed as fluorescence versus dose, e.g., in anormal dose-response fashion. The dose levels that result in the upperplateau of the curve represent the levels needed to obtain efficacy inan in vivo model.

[0386] This assay may also be used to determine the plasma levels neededto saturate the binding sites of other integrins, such as the α₉β₁integrin, which is the integrin most closely related α₄β₁ (Palmer et al,1993, J. Cell Bio., 123:1289). Such binding is predictive of in vivoutility for inflammatory conditions mediated by α₉β₁ integrin, includingby way of example, airway hyper-responsiveness and occlusion that occurswith chronic asthma, smooth muscle cell proliferation inatherosclerosis, vascular occlusion following angioplasty, fibrosis andglomerular scarring as a result of renal disease, aortic stenosis,hypertrophy of synovial membranes in rheumatoid arthritis, andinflammation and scarring that occur with the progression of ulcerativecolitis and Crohn's disease.

[0387] Accordingly, the above-described assay may be performed with ahuman colon carcinoma cell line, SW 480 (ATTC #CCL228) transfected withcDNA encoding α₉ integrin (Yokosaki et al., 1994, J. Biol. Chem.,269:26691), in place of the Jurkat cells, to measure the binding of theα₉β₁ integrin. As a control, SW 480 cells which express other α and β₁subunits may be used.

[0388] Accordingly, another aspect of this invention is directed to amethod for treating a disease in a mammalian patient, which disease ismediated by α₉β₁, and which method comprises administering to saidpatient a therapeutically effective amount of a compound of thisinvention. Such compounds are preferably administered in apharmaceutical composition described herein above. Effective dailydosing will depend upon the age, weight, condition of the patient whichfactors can be readily ascertained by the attending clinician. However,in a preferred embodiment, the compounds are administered from about 20to 500 μg/kg per day.

Example C In vivo Evaluation

[0389] The standard multiple sclerosis model, Experimental Autoimmune(or Allergic) Encephalomyelitis (“EAE”), was used to determine theeffect of candidate compounds to reduce motor impairment in rats orguinea pigs. Reduction in motor impairment is based on blocking adhesionbetween leukocytes and the endothelium and correlates withanti-inflammatory activity in the candidate compound. This model hasbeen previously described by Keszthelyi et al., Neurology, 1996,47:1053-1059, and measures the delay of onset of disease.

[0390] Brains and spinal cords of adult Hartley guinea pigs werehomogenized in an equal volume of phosphate-buffered saline. An equalvolume of Freund's complete adjuvant (100 mg mycobacterium tuberculosisplus 10 ml Freund's incomplete adjuvant) was added to the homogenate.The mixture was emulsified by circulating it repeatedly through a 20 mlsyringe with a peristaltic pump for about 20 minutes.

[0391] Female Lewis rats (2-3 months old, 170-220 g) or Hartley guineapigs (20 day old, 180-200 g) were anesthetized with isoflurane and threeinjections of the emulsion, 0.1 ml each, were made in each flank. Motorimpairment onset is seen in approximately 9 days.

[0392] Candidate compound treatment began on Day 8, just before onset ofsymptoms. Compounds were administered subcutaneously (“SC”), orally(“PO”) or intraperitoneally (“IP”). Doses were given in a range of 10mg/kg to 200 mg/kg, bid, for five days, with typical dosing of 10 to 100mg/kg SC, 10 to 50 mg/kg PO, and 10 to 100 mg/kg IP.

[0393] Antibody GG5/3 against α₄β₁ integrin (Keszthelyi et al.,Neurology, 1996, 47:1053-1059), which delays the onset of symptoms, wasused as a positive control and was injected subcutaneously at 3 mg/kg onDay 8 and 11.

[0394] Body weight and motor impairment were measured daily. Motorimpairment was rated with the following clinical score: 0 no change 1tail weakness or paralysis 2 hindlimb weakness 3 hindlimb paralysis 4moribund or dead

[0395] A candidate compound was considered active if it delayed theonset of symptoms, e.g., produced clinical scores no greater than 2 orslowed body weight loss as compared to the control.

Example D Asthma Model

[0396] Inflammatory conditions mediated by α₄β₁ integrin include, forexample, airway hyper-responsiveness and occlusion that occurs withchronic asthma. The following describes an asthma model which can beused to study the in vivo effects of the compounds of this invention foruse in treating asthma.

[0397] Following the procedures described by Abraham et al, J. Clin.Invest, 93:776-787 (1994) and Abraham et al, Am J. Respir Crit Care Med,156:696-703 (1997), both of which are incorporated by reference in theirentirety. Compounds of this invention are formulated into an aerosol andadministered to sheep which are hypersensitive to Ascaris suum antigen.Compounds which decrease the early antigen-induced bronchial responseand/or block the late-phase airway response, e.g., have a protectiveeffect against antigen-induced late responses and airwayhyper-responsiveness (“AHR”), are considered to be active in this model.

[0398] Allergic sheep which are shown to develop both early and latebronchial responses to inhaled Ascaris suum antigen are used to studythe airway effects of the candidate compounds. Following topicalanesthesia of the nasal passages with 2% lidocaine, a balloon catheteris advanced through one nostril into the lower esophagus. The animalsare then intubated with a cuffed endotracheal tube through the othernostril with a flexible fiberoptic bronchoscope as a guide.

[0399] Pleural pressure is estimated according to Abraham (1994).Aerosols (see formulation below) are generated using a disposablemedical nebulizer that provides an aerosol with a mass medianaerodynamic diameter of 3.2 μm as determined with an Andersen cascadeimpactor. The nebulizer is connected to a dosimeter system consisting ofa solenoid valve and a source of compressed air (20 psi). The output ofthe nebulizer is directed into a plastic T-piece, one end of which isconnected to the inspiratory port of a piston respirator. The solenoidvalve is activated for 1 second at the beginning of the inspiratorycycle of the respirator. Aerosols are delivered at V_(T) of 500 ml and arate of 20 breaths/minute. A 0.5% sodium bicarbonate solution only isused as a control.

[0400] To assess bronchial responsiveness, cumulativeconcentration-response curves to carbachol can be generated according toAbraham (1994). Bronchial biopsies can be taken prior to and followingthe initiation of treatment and 24 hours after antigen challenge.Bronchial biopsies can bepreformed according to Abraham (1994).

[0401] An in vitro adhesion study of alveolar macrophages can also beperformed according to Abraham (1994), and a percentage of adherentcells is calculated.

[0402] Aerosol Formulation

[0403] A solution of the candidate compound in 0.5% sodiumbicarbonate/saline (w/v) at a concentration of 30.0 mg/mL is preparedusing the following procedure:

[0404] A. Preparation of 0.5% Sodium Bicarbonate/Saline Stock Solution:100.0 mL Ingredient Gram/100.0 mL Final Concentration Sodium Bicarbonate0.5 g 0.5% Saline q.s. ad 100.0 mL q.s. ad 100%

[0405] Procedure:

[0406] 1. Add 0.5 g sodium bicarbonate into a 100 mL volumetric flask.

[0407] 2. Add approximately 90.0 mL saline and sonicate until dissolved.

[0408] 3. Q.S. to 100.0 mL with saline and mix thoroughly.

[0409] B. Preparation of 30.0 mg/mL Candidate Compound: 10.0 mLIngredient Gram/10.0 mL Final Concentration Candidate Compound 0.300 g30.0 mg/mL 0.5% Sodium q.s. ad 10.0 mL q.s ad 100% Bicarbonate/SalineStock Solution

[0410] Procedure:

[0411] 1. Add 0.300 g of the candidate compound into a 10.0 mLvolumetric flask.

[0412] 2. Add approximately 9.7 mL of 0.5% sodium bicarbonate/salinestock solution.

[0413] 3. Sonicate until the candidate compound is completely dissolved.

[0414] 4. Q.S. to 10.0 mL with 0.5% sodium bicarbonate/saline stocksolution and mix thoroughly.

[0415] Using a conventional oral formulation, compounds of thisinvention would be active in this model.

Example E Allograft Model

[0416] Allograft rejection, associated with infiltration of inflammatorycells, is the leading obstacle to long-term allograft survival. Cellsurface adhesion molecules facilitate alloantigen recognition in vitroand may be critical for lymphocyte traffic in vivo. The followingdescribes a model which can be used to study the in vivo effects of thecompounds of this invention in the control of allograft rejection.

[0417] The following procedures are described in Coito et al.,Transplantation (1998) 65(6):699-706 and in Korom et al.,Transplantation (1998) 65(6):854-859, both of which are incorporated byreference in their entirety.

[0418] Following the procedures described in Coito and Korom, male adultrats weighing approximately 200-250 g are used in this model. Lewis ratsare used as the recipients of cardiac allografts from Lewis X BrownNorway rats. Hearts are transplanted into the abdominal great vesselsusing standard microvascular techniques.

[0419] A candidate compound is administered to the transplant recipientin a suitable pharmaceutical carrier for a 7-day course of treatmentstarting the day of the engraftment. Doses range from 0.3 to 30mg/kg/day. Control recipients receive the pharmaceutical carrier only.The rats are euthanized and their cardiac allografts are analyzed asdescribed in Coito and Korom.

[0420] Using conventional formulations, compounds of this inventionwould be active in this model.

1-10. (cancelled)
 11. The compound of formula IId;

wherein: HetAr is a nitrogen containing heteroaryl group substituted with a substituent selected from the group consisting of acyl, acylamino, acyloxy, aminoacyl, aminocarbonylamino, aminothiocarbonylamino, aminocarbonyloxy, oxycarbonylamino, oxythiocarbonylamino, thioamidino, thiocarbonylamino, aminosulfonylamino, aminosulfonyloxy, aminosulfonyl, oxysulfonylamino, aryl, substituted aryl, and oxysulfonyl; R¹⁶ and R¹⁷ are independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkoxy, substituted alkoxy, amino, substituted amino, cycloalkyl, substituted cycloalkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic, substituted heterocyclic and halogen; R²¹ is selected from the group consisting of alkyl, substituted alkyl, alkoxy, substituted alkoxy, amino, substituted amino, cycloalkyl, substituted cycloalkyl, aryl, substituted aryl, heterocyclic and substituted heterocyclic; and X is hydroxyl; and enantiomers, diastereomers or pharmaceutically acceptable salts thereof.
 12. (cancelled)
 13. The compound of claim 11 wherein HetAr is a nitrogen containing heteroaryl group which is substituted with a group of formula —O-Z-NR¹¹R^(11′) or —O-Z-R¹² wherein R¹¹ and R^(11′) are independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, heterocyclic, substituted heterocyclic, and where R¹¹ and R^(11′) are joined to form a heterocycle or a substituted heterocycle, R¹² is selected from the group consisting of heterocycle and substituted heterocycle, and Z is selected from the group consisting of —C(O)— and —SO₂—.
 14. The compound of claim 13 wherein the nitrogen containing heteroaryl group is substituted with a group of formula —OC(O)NR¹¹R^(11′) wherein R¹¹ and R^(11′) are independently selected from the group consisting of alkyl or R¹¹ and R^(11′) are joined to form a heterocycle or a substituted heterocycle.
 15. The compound of claim 14 wherein the nitrogen containing heteroaryl group is substituted with —OC(O)N(CH₃)₂ and is at the para position of the heteroaryl group.
 16. The compound of claim 11 wherein HetAr is a nitrogen containing heteroaryl group which is substituted with an aryl or substituted aryl group.
 17. (cancelled)
 18. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and a therapeutically effective amount of a compound of claim
 11. 19. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and a therapeutically effective amount of a compound of claim
 13. 20. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and a therapeutically effective amount of a compound of claim
 14. 21. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and a therapeutically effective amount of a compound of claim
 15. 22. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and a therapeutically effective amount of a compound of claim
 16. 